Anti-ly6k antibody-drug conjugates and methods of use

Abstract

The present disclosure provides antibody-drug conjugates (ADCs) that specifically bind to lymphocyte antigen 6 family member K (LY6K). Also provided are compositions comprising these ADCs and methods of treating a subject using these ADCs.

Description

ANTLLY6K ANTIBODY-DRUG CONJUGATES AND METHODS OF USECROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit of and priority to United States Provisional Application Serial Number 63 / 709,771, filed October 21, 2024, the contents of which are hereby incorporated by reference herein in their entirety.REFERENCE TO SEQUENCE LISTING

[0002] This application contains a sequence listing which has been submitted electronically in ST.26 format and is hereby incorporated by reference in its entirety. Said ST.26 copy, created on October 21, 2025, is named “222644 seqlist.xml” and is 223,261 bytes in size.FIELD

[0003] The present disclosure relates to antibody-drug conjugates that are specific for human lymphocyte antigen 6 family member K (LY6K), and methods of use thereof.BACKGROUND

[0004] The lymphocyte antigen 6 (LY6) family is a group of cysteine-rich proteins that share a common structure, are expressed in various tissues, and exhibit a wide variety of cellular functions. LY6 proteins are anchored to the cell membrane via glycosylphosphatidylinositol (GPI) anchor or secreted, and they are involved in cell proliferation, cell migration, cell-cell interactions, immune cell maturation, macrophage activation, and cytokine production. Dysregulated expression of LY6 family members is associated with tumorigenesis and autoimmune diseases, and many LY6 proteins are used as tumor markers and as potential therapeutic targets. Lymphocyte antigen 6 family member K (LY6K) is a cell surface protein that is upregulated in numerous types of cancer, including esophageal squamous cell carcinoma, bladder cancer, and breast cancer, and contributes to cell growth, migration, invasion, and immune escape.

[0005] Thus, there is a need for therapies targeting LY6K.SUMMARY

[0006] The present disclosure provides antibody-drug conjugates (ADCs) that specifically bind to LY6K (e.g., human LY6K). Also provided are pharmaceutical compositions comprising these ADCs and methods of treating a subject using these ADCs. The ADCs provided herein areparticularly advantageous because they specifically bind LY6K-expressing cancer cells, allowing targeted delivery of therapeutic and diagnostic agents, and therefore have utility in the treatment and diagnosis of cancer (e., LY6K-expressing cancer).

[0007] In one aspect, provided herein is an antibody-drug conjugate (ADC) according to general structure (1):AB-[(L6)b-{Z-L-D}x]y(1)wherein:AB is an antibody that specifically binds human LY6K;L is a linker that links Z to D;Z is a connecting group;L6is -GlcNAc(Fuc)w-(G)j-S-(L7)W’-, wherein G is a monosaccharide, j is an integer in the range of 0 - 10, S is a sugar or a sugar derivative, GlcNAc is N-acetylglucosamine and Fuc is fucose, w is 0 or 1, w’ is 0, 1 or 2 and L7is -N(H)C(O)CH2-, -N(H)C(O)CF2- or -CH2-;D is selected from the group consisting of taxanes, anthracyclines, camptothecins, epothilones, mytomycins, combretastatins, vinca alkaloids, tubulysins, enediynes, amanitins, duocarmycins, maytansinoids, amatoxins, dolastatins, auristatins, eribulins, BCL-XL inhibitors, hemiasterlins, KSP inhibitors, radioisotopes, TLR agonists, indolinobenzodiazepine dimers or pyrrolobenzodiazepine dimers (PBDs), and analogues or prodrugs thereof;b is 0 or 1;x is 1 or 2; andy is 1, 2, 3 or 4.

[0008] In certain embodiments, linker L has the structure-(L’ ^p-CLwherein:L1, L2, L3and L4are each individually linkers that together link Z to D;n, o, p and q are each individually 0 or 1, provided that n + o + p + q= l, 2, 3 or 4, optionally wherein n = 0 = p = 1;and wherein:(a) linker L1is represented by:-(W)k-(A)d-(B)e-(A)f-(B)g-(C(O))g-, or-(W)k-(A)d-(B)e-(A)f-(C(O))g-N*[-(A)d’-(B)e’-(A)f-(C(O))g-]2, wherein:d and d’ are individually 0 or 1;e and e’ are individually an integer in the range 1 - 10;f and f are individually 0, or 1;g and g’ are individually an integer in the range 0 - 10;k = 0 or 1 with the proviso that if k = 1 then d = 0;A is a sulfamide group according to structure (23)wherein a = 0 or 1, and R13is selected from the group consisting of hydrogen, Ci - C24 alkyl groups, C3 - C24 cycloalkyl groups, C2 - C24 (hetero)aryl groups, C3 - C24 alkyl(hetero)aryl groups and C3 - C24 (hetero)arylalkyl groups, the Ci - C24 alkyl groups, C3 - C24 cycloalkyl groups, C2 - C24 (hetero)aryl groups, C3 - C24 alkyl(hetero)aryl groups and C3 - C24 (hetero)aryl alkyl groups optionally substituted and optionally interrupted by one or more heteroatoms selected from O, S and NR14wherein R14is independently selected from the group consisting of hydrogen and Ci - C4 alkyl groups, or R13is D connected to N via a spacer moiety, optionally wherein the spacer moiety is -(B)g-(C(O))g-(L2)o-(L3)p-(L4)q-;W is -OC(O)-, -C(O)O- -C(O)NH- -NHC(O)-, -OC(O)NH-, -NHC(O)O- -C(O)(CH2)mC(O)-, -C(O)(CH2)raC(O)NH- or -(4-Ph)CH2NHC(O)(CH2)mC(O)NH- wherein m is an integer in the range 0 - 10;B is a -CH2-CH2-O- or a -O-CH2-CH2- moiety, or (B)eis a -(CH2-CH2-O)ei-CH2- CH2- moiety, wherein el is an integer in the range 1 - 10;N* is a branching nitrogen atom, to which two instances of-(A)d-(B)e-(A)f-(C(O))g’- are connected and both (C(O))g’ moieties are connected to -(L2)0-(L3)p-(L4)q-D, wherein L2, L3, L4, o, p, q and D are each selected individually;and / or(b) linker L2is a peptide spacer, optionally wherein linker L2comprises 1 - 5 amino acids, wherein linker L2is a dipeptide, tripeptide or tetrapeptide spacer, or wherein linker L2isrepresented by general structure (L3):wherein R17= CH3or CH2CH2CH2NHC(O)NH2;and / or(c) linker L3is a self-immolative spacer, optionally wherein linker L3is a paraaminobenzyloxycarbonyl (PABC) derivative according to structure (L4).wherein R21is H, R26or C(O) R26, wherein R26is Ci - C24 (hetero)alkyl groups, C3- Cio (hetero)cycloalkyl groups, C2- Cio (hetero)aryl groups, C3- Cio alkyl(hetero)aryl groups and C3- Cio (hetero)arylalkyl groups, which are optionally substituted and optionally interrupted by one or more heteroatoms selected from O, S and NR28, wherein R28is independently selected from the group consisting of hydrogen and Ci - C4 alkyl groups,optionally wherein R21is H or C(O)R26, wherein R26= 4-methyl-piperazine or morpholine;and / or(d) linker L4is an aminoalkanoic acid spacer according to the structure - NR22-(Cz-alkylene)- C(O)-, wherein x is an integer in the range 1 - 20 and R22is H or Ci - C4 alkyl; or linker L4is a an ethyleneglycol spacer according to the structure -NR22-(CH2-CH2-O)e6- (CH2)e7-C(O)-, wherein e6 is an integer in the range 1 - 10, e7 is an integer in the range 1 - 3 and R22is H or Ci - C4 alkyl; orlinker L4is a diamine spacer according to the structure -NR22-(Cz-alkylene)-NR22- (C(O))h-, wherein h is 0 or 1, x is an integer in the range 1 - 10 and R22is H or Ci - C4 alkyl.

[0009] In certain embodiments, b = 1 and conjugation is via the glycan of the antibody, optionally wherein j = 0.

[0010] In certain embodiments, L-D has a structure selected from the group consisting of (X) - (XIII):(XIII)wherein the wavy line indicates the connection to Z, and L2, L4, o and q are as defined in claim 2.

[0011] In certain embodiments, L-D has a structure selected from the group consisting of (Xa), (Xlb), (Xllg), (Xllle) and (Xllh):wherein the wavy line indicates the connection to Z, and R22is as defined above.

[0012] In certain embodiments, Z comprises a cyclooctenyl group and is according to structure (Z42):wherein:the bond labelled with * is connected to S and the wavy bond labelled with ** is connected to L;each R15is independently selected from the group consisting of hydrogen, halogen, - OR16, -NO2, -CN, -S(O)2R16, -S(O)3('), CI - C24 alkyl groups, C5 - C24 (hetero)aryl groups, C7 - C24 alkyl(hetero)aryl groups and C7 - C24 (hetero)aryl alkyl groups and wherein the alkyl groups, (hetero)aryl groups, alkyl(hetero)aryl groups and (hetero)arylalkyl groups are optionally substituted, wherein two substituents R15may be linked together to form an optionally substituted annulated cycloalkyl or an optionally substituted annulated (hetero)arene substituent, and wherein R16is independently selected from the group consisting of hydrogen, halogen, Ci - C24 alkyl groups, Ce - C24 (hetero)aryl groups, C7 - C24 alkyl(hetero)aryl groups and C7 - C24 (hetero)aryl alkyl groups;each R18is independently selected from the group consisting of hydrogen, halogen, Ci - C24 alkyl groups, Ce - C24 (hetero)aryl groups, C7 - C24 alkyl(hetero)aryl groups and C7 - C24 (hetero)arylalkyl groups;R19is selected from the group consisting of hydrogen, halogen, Ci - C24 alkyl groups, Ce - C24 (hetero)aryl groups, C7 - C24 alkyl(hetero)aryl groups and C7 - C24 (hetero)arylalkyl groups, the alkyl groups optionally being interrupted by one of more hetero-atoms selected from the group consisting of O, N and S, wherein the alkyl groups, (hetero)aryl groups, alkyl(hetero)aryl groups and (hetero)arylalkyl groups are independently optionally substituted, or R19is a second occurrence of Z (or Q) or D connected via a spacer moiety; and1 is an integer in the range 0 to 10.

[0013] In another aspect, provided herein is an antibody-drug conjugate (ADC) according to general structure (1):AB-[(L6)b-{Z-L-D}x]y(1)wherein the ADC is obtained by a process comprising:(i) contacting an antibody that specifically binds human LY6K comprising y core N-acetylglucosamine (GlcNAc) moi eties, wherein y = 1, 2, 3 or 4, with a compound of the formula S(F)X-P in the presence of a catalyst, wherein S(F)Xis a sugar derivative comprising x reactive groups F capable of reacting with a reactive group Q, x is 1 or 2 and P is a nucleoside mono- ordiphosphate, and wherein the catalyst is capable of transferring the S(F)Xmoiety to the core-GlcNAc moiety, to obtain a modified antibody according to Formula (26):AB-[GlcNAc(Fuc)w-S {F }x]y(26)whereinAB is the antibody that specifically binds human LY6K;Fuc is fucose; andw is 0 or 1; and(ii) reacting the modified antibody with a compound according to structure (2):Q-L-D(2)wherein:Q is a reactive moiety;L is a linker that links Z to D; andD is selected from the group consisting of taxanes, anthracyclines, camptothecins, epothilones, mytomycins, combretastatins, vinca alkaloids, tubulysins, enediynes, amanitins, duocarmycins, maytansinoids, amatoxins, dolastatins, auristatins, eribulins, BCL-XL inhibitors, hemiasterlins, KSP inhibitors, radioisotopes, TLR agonists, indolinobenzodiazepine dimers or pyrrolobenzodiazepine dimers (PBDs), and analogues or prodrugs thereof;to obtain the ADC.

[0014] In certain embodiments of any of the ADCs provided herein the antibody that specifically binds human LY6K comprises: a VH comprising the CDRH1, CDRH2, and CDRH3 amino acid sequences of the VH amino acid sequence set forth in any one of SEQ ID NOs: 1-34; and a VL comprising the CDRL1, CDRL2, and CDRL3 amino acid sequences of the VL amino acid sequence set forth in any one of SEQ ID NOs: 35-68.

[0015] In certain embodiments, the VH amino acid sequence and VL amino acid sequence are as set forth in: SEQ ID NOs: 1 and 35, 2 and 36, 3 and 37, 4 and 38, 5 and 39, 6 and 40, 7 and 41, 8 and 42, 9 and 43, 10 and 44, 11 and 45, 12 and 46, 13 and 47, 14 and 48, 15 and 49, 16 and 50, 17 and 51, 18 and 52, 19 and 53, 20 and 54, 21 and 55, 22 and 56, 23 and 57, 24 and 58, 25 and 59, 26 and 60, 27 and 61, 28 and 62, 29 and 63, 30 and 64, 31 and 65, 32 and 66, 33 and 67, or 34 and 68, respectively.

[0016] In certain embodiments, the antibody comprises the CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in: SEQ ID NOs: 69, 90, and 124; 70, 91, and 125; 71, 92, and 126; 72, 93, and 127; 69, 94, and 124; 73, 95, and 128; 74, 96, and 129; 75, 97, and 130; 74, 98, and 131; 76, 99, and 132; 77, 100, and 133; 78, 101, and 134; 79, 102, and 135; 74, 103, and 136; 80, 104, and 137; 74, 105, and 138; 74, 106, and 139; 81, 107, and 140; 81, 108, and 141; 82, 109, and 142; 83, 110, and 143; 84, 99, and 144; 85, 111, and 145; 81, 112, and 146; 86, 113, and 147; 87, 114, and 148; 74, 115, and 149; 74, 116, and 150; 74, 117, and 151; 88, 118, and 152; 89, 119, and 153; 81, 120, and 154; 74, 121, and 155; 85, 122, and 156; or 86, 123, and 157, respectively.

[0017] In certain embodiments, the antibody comprises the CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in: SEQ ID NO: 158, KIS, and SEQ ID NO: 198; SEQ ID NO: 159, WAS, and SEQ ID NO: 199; SEQ ID NO: 160, STT, and SEQ ID NO: 200; SEQ ID NO: 161, WAS, and SEQ ID NO: 201; SEQ ID NO: 162, AAS, and SEQ ID NO: 202; SEQ ID NOs: 163, 185, and 203; SEQ ID NOs: 164, 185, and 204; SEQ ID NOs: 165, 185, and 205; SEQ ID NOs: 166, 186, and 206; SEQ ID NOs: 167, 187, and 207; SEQ ID NOs: 168, 188, and 208; SEQ ID NOs: 169, 186, and 209; SEQ ID NOs: 170, 185, and 198; SEQ ID NOs: 169, 189, and 210; SEQ ID NOs: 170, 185, and 211; SEQ ID NOs: 171, 187, and 212; SEQ ID NOs: 172, 187, and 213; SEQ ID NOs: 173, 190, and 214; SEQ ID NOs: 174, 191, and 215; SEQ ID NOs: 175, 186, and 216; SEQ ID NOs: 176, 186, and 217; SEQ ID NOs: 170, 192, and 198; SEQ ID NOs: 171, 193, and 218; SEQ ID NOs: 177, 187, and 219; SEQ ID NOs: 178, 189, and 220; SEQ ID NOs: 179, 187, and 221; SEQ ID NOs: 165, 185, and 222; SEQ ID NOs: 180, 187, and 223; SEQ ID NOs: 181, 194, and 224; SEQ ID NOs: 182, 195, and 199; SEQ ID NOs: 172, 187, and 225; SEQ ID NOs: 183, 196, and 226; or SEQ ID NOs: 184, 197, and 227, respectively.

[0018] In certain embodiments, the antibody comprises the CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in: SEQ ID NOs: 69, 90, 124, 158, KIS, and SEQ ID NO: 198; SEQ ID NOs: 70, 91, 125, 159, WAS, and SEQ ID NO: 199; SEQ ID NOs: 71, 92, 126, 160, STT, and SEQ ID NO: 200; SEQ ID NOs: 72, 93, 127, 161, WAS, and SEQ ID NO: 201; SEQ ID NOs: 69, 94, 124, 158, KIS, and SEQ ID NO: 198; SEQ ID NOs: 73, 95, 128, 162, AAS, and SEQ ID NO: 202; SEQ ID NOs: 74, 96, 129, 163, 185, and 203; SEQ ID NOs: 75, 97, 130, 164, 185, and 204; SEQ ID NOs: 74, 98, 131, 165, 185, and 205; SEQ ID NOs: 76, 99, 132, 166, 186, and 206; SEQ ID NOs: 77, 100, 133, 167, 187, and 207; SEQ ID NOs: 78, 101, 134, 168, 188, and 208; SEQ ID NOs: 79, 102, 135, 169, 186, and 209; SEQ ID NOs: 74,103, 136, 170, 185, and 198; SEQ ID NOs: 80, 104, 137, 169, 189, and 210; SEQ ID NOs: 74, 105, 138, 170, 185, and 211; SEQ ID NOs: 74, 106, 139, 171, 187, and 212; SEQ ID NOs: 81, 107, 140, 172, 187, and 213; SEQ ID NOs: 81, 108, 141, 173, 190, and 214; SEQ ID NOs: 82, 109, 142, 174, 191, and 215; SEQ ID NOs: 83, 110, 143, 175, 186, and 216; SEQ ID NOs: 84, 99, 144, 176, 186, and 217; SEQ ID NOs: 85, 111, 145, 170, 192, and 198; SEQ ID NOs: 81, 112, 146, 171, 193, and 218; SEQ ID NOs: 86, 113, 147, 177, 187, and 219; SEQ ID NOs: 87, 114, 148, 178, 189, and 220; SEQ ID NOs: 74, 115, 149, 179, 187, and 221; SEQ ID NOs: 74, 116, 150, 165, 185, and 222; SEQ ID NOs: 74, 117, 151, 180, 187, and 223; SEQ ID NOs: 88, 118, 152, 181, 194, and 224; SEQ ID NOs: 89, 119, 153, 182, 195, and 199; SEQ ID NOs: 81, 120, 154, 172, 187, and 225; SEQ ID NOs: 74, 121, 155, 165, 185, and 222; SEQ ID NOs: 85, 122, 156, 183, 196, and 226; or SEQ ID NOs: 86, 123, 157, 184, 197, and 227, respectively.

[0019] In certain embodiments, the antibody comprises the VH amino acid sequence set forth in any one of SEQ ID NOs: 1-34.

[0020] In certain embodiments, the antibody comprises a heavy chain constant region, optionally selected from the group consisting of human IgGl, IgG2, IgG3, IgG4, IgAl, and IgA2.

[0021] In certain embodiments, the antibody comprises a heavy chain constant region that is a variant of a wild-type heavy chain constant region, wherein the variant heavy chain constant region binds to an FcyR with lower affinity than the wild-type heavy chain constant region binds to the FcyR.

[0022] In certain embodiments, the antibody comprises a heavy chain constant region comprising the amino acid sequence of SEQ ID NO: 228 or 229.

[0023] In certain embodiments, the antibody comprises the VL amino acid sequence set forth in any one of SEQ ID NOs: 35-68.

[0024] In certain embodiments, the antibody comprises a light chain constant region comprising the amino acid sequence of SEQ ID NO: 230 or 231.

[0025] In certain embodiments, the VH and VL comprise the amino acid sequences set forth in: SEQ ID NOs: 1 and 35, 2 and 36, 3 and 37, 4 and 38, 5 and 39, 6 and 40, 7 and 41, 8 and 42, 9 and 43, 10 and 44, 11 and 45, 12 and 46, 13 and 47, 14 and 48, 15 and 49, 16 and 50, 17 and 51, 18 and 52, 19 and 53, 20 and 54, 21 and 55, 22 and 56, 23 and 57, 24 and 58, 25 and 59, 26 and 60, 27 and 61, 28 and 62, 29 and 63, 30 and 64, 31 and 65, 32 and 66, 33 and 67, or 34 and 68, respectively, respectively.

[0026] In certain embodiments, the antibody specifically binds to an amino acid sequence selected from the group consisting of SEQ ID NOs: 233-238.

[0027] In certain embodiments, the antibody specifically binds to the amino acid sequence of SEQ ID NOs: 233 and 239.

[0028] In certain embodiments, the antibody specifically binds to the amino acid sequence of SEQ ID NOs: 234 and 240.

[0029] In certain embodiments, the antibody specifically binds to the amino acid sequence of SEQ ID NOs: 237 and 247.

[0030] In certain embodiments, D is selected from the group consisting of anthracyclines, camptothecins, maytansinoids, enediynes, auristatins and pyrrolobenzodiazepine dimers.

[0031] In certain embodiments, D is an auristatin selected from the group consisting of MMAD, MMAE, MMAF and PF-06380101. In certain embodiments, is MMAE or PF-06380101. In certain embodiments, wherein D is MMAE.

[0032] In certain embodiments, D is a camptothecin selected from the group consisting of topotecan, silatecan, cositecan, exatecan, exatecan-S, DXd, SN-38, lurtotecan, gimatecan, belotecan, rubitecan, AMDCPT, and G-AMDCPT. In certain embodiments, D is exatecan or DXd. In certain embodiments, D is exatecan.

[0033] In certain embodiments, D is an enediyne selected from the group consisting of calicheamicins, esperamicins, shishijimicins, and namenamicins. In certain embodiments, D is calicheamicin.

[0034] In another aspect, provided herein is a composition comprising an ADC described herein and a pharmaceutically acceptable carrier or excipient.

[0035] In another aspect, provided herein is a method of treating cancer in a subject, the method comprising administering to the subject an effective amount of an ADC or composition described herein. In certain embodiments, the method further comprises administering an additional therapeutic agent to the subject. In certain embodiments, the additional therapeutic agent is a chemotherapeutic, a radiotherapeutic, or a checkpoint targeting agent. In certain embodiments, the checkpoint targeting agent is selected from the group consisting of an antagonist anti-PD-1 antibody, an antagonist anti-PD-Ll antibody, an antagonist anti-PD-L2 antibody, an antagonist anti-CTLA-4 antibody, an antagonist anti-TIM-3 antibody, an antagonist anti-LAG-3 antibody, an antagonist anti-VISTA antibody, an antagonist anti-TIGIT antibody, an antagonist anti-CD96antibody, an antagonist anti-CEACAMl antibody, an agonist anti-CD137 antibody, an agonist anti-GITR antibody, and an agonist anti-OX40 antibody.

[0036] In another aspect, provided herein is a use of an ADC or composition described herein for the manufacture of a medicament for treating cancer.

[0037] In another aspect, provided herein is an ADC or composition described herein for use in medicine.

[0038] In another aspect, provided herein is an ADC or composition described herein for use in treating cancer in a subject.BRIEF DESCRIPTION OF THE DRAWINGS

[0039] FIG. 1 shows a representative set of reactive groups (F) that when present in a biomolecule lead to connecting group Z (la-lh) upon reaction with reactive group Q, according to aspects of the present disclosure. Functional groups F can be naturally present or may be artificially introduced (engineered) into a biomolecule (e., antibody) at any position of choice.

[0040] FIGs. 2A-2F show structures of exemplary ADCs provided herein, according to aspects of the present disclosure.

[0041] FIGs. 3A and 3B show structures of exemplary compounds according to general structure (2), according to aspects of the present disclosure.

[0042] FIG. 4 shows that exemplary anti-LY6K antibodies provided herein selectively bind LY6K, according to aspects of the present disclosure. Depicted are the plate reading results of ELISA assays measuring the binding of the indicated anti-LY6K antibodies to LY6K, LY6E, LY6D, LY6H, and BSA (negative control).

[0043] FIG. 5 shows that exemplary anti-LY6K antibodies provided herein detect LY6K, according to aspects of the present disclosure. Depicted in the left panel are the results of a capture ELISA measuring the sensitivity of the indicated anti-LY6K antibodies, as well as positive control commercially available anti-LY6K antibodies from R& D Systems and Abeam, for detecting LY6K. The table in the right panel indicates the sensitivity and half maximal effective concentration (EC50) for each of the indicated antibodies.

[0044] FIG. 6 shows that an anti-LY6K antibody provided herein is internalized into cells, according to aspects of the present disclosure. Depicted is a representative fluorescence plot for anti-LY6K antibody 3082 labeled with pH-sensitive dye. Upon internalization into OVCAR-8 cells, the antibody is endocytosed and releases fluorescent signal. Also depicted are results fornegative control huIgGl and positive control trastuzumab, a known commercially available internalizer.

[0045] FIG. 7 shows internalization of exemplary anti-LY6K antibodies provided herein, according to aspects of the present disclosure. Data depicted are a representative fluorescence plot for anti-LY6K antibodies 2660 & 3481 labeled with pH-sensitive dye. Upon internalization into OVCAR-8 cells, the labeled antibody is endocytosed, internalized into the acidic environment of the lysosome, and releases fluorescent signal. The area of fluorescence and intensity inside the cell increases. This is measured using a live-cell analysis system for image based fluorescent measurements of antibody internalization, which analyzes the change in fluorescence intensity over time. Also depicted are results for negative control human IgGl and positive control trastuzumab, a known commercially available internalizer.

[0046] FIGs. 8A and 8B show increased cytotoxicity in LY6K-expressing cells upon treatment with anti-LY6K ADCs, according to aspects of the present disclosure. SYNstatin ADCs (FIG. 8A) or SYNtecan ADCs (FIG. 8B) made with the 2660 anti-LY6K antibody or the 3481 anti-LY6K antibody were added to wells with HEK-293-LY6K over expressing cells in the diluation range of 50-0.2 pg / ml (337-1.3 nM), along with ADCs made using the isotype control antibody. Cell viability was evaluated 4 days after the treatment using CellTiter-Glo® Luminescent Cell Viability Assay (Promega) and a microplate reader according to manufacturer’s instructions. All readings were normalized to the percentage of the value derived from the control untreated wells and EC50 values were calculated by a non-linear regression curve in GraphPad Prism.

[0047] FIGs. 9A and 9B show reduction in tumor volume in a lung cancer model in mice upon treatment with anti-LY6K ADCs, according to aspects of the present disclosure. SYNstatin ADCs made with the 2660 anti-LY6K antibody (FIG. 9A) or the 3481 anti-LY6K antibody (FIG. 9B) were dosed weekly starting at day 7 post tumor inoculation. Tumor volumes were measured by digital caliper twice a week. Data shown are mean tumor volumes ± SEM of 10 mice per ADC treatment group or 5 mice for the vehicle control group.

[0048] FIGs. 10A and 10B show reduction in tumor volume in a breast cancer model in mice upon treatment with anti-LY6K ADCs, according to aspects of the present disclosure. SYNtecan ADCs made with the 2660 anti-LY6K antibody (FIG.10A) or the 3481 anti-LY6K antibody (FIG.10B) were dosed on days 14, 35, and 42 post tumor inoculation. Tumor volumes were measuredby digital caliper twice a week. Data shown are mean tumor volumes ± SEM of 10 mice per ADC treatment group or 5 mice for the vehicle control group.DETAILED DESCRIPTION

[0049] The instant disclosure provides anti-LY6K antibody-drug conjugates (ADCs). Also provided are pharmaceutical compositions comprising these ADCs and methods of treating a subject using these ADCs. The ADCs disclosed herein are particularly useful for diagnosing and / or treating cancer in a subj ect.Definitions

[0050] As used herein, the term “LY6K” refers to lymphocyte antigen 6 family member K, a membrane-anchored protein that is highly expressed in cancer cells and tissues. As used herein, the term “human LY6K” refers to a protein encoded by a wild-type human LY6K gene (e.g., the gene set forth in NM 017527.4). The amino acid sequence of an exemplary human LY6K protein (as set forth in RefSeq NP_059997.3) is MALLALLLVVALPRVWTDANLTARQRDPEDSQRTDEGDNRVWCHVCERENTFEC QNPRRCKWTEPYCVIAAVKIFPRFFMVAKQCSAGCAAMERPKPEEKRFLLEEPMPF FYLKCCKIRYCNLEGPPINSSVFKEYAGSMGESCGGLWLAILLLLASIAAGLSLS(SEQ ID NO: 232).

[0051] As used herein, the terms “antibody” and “antibodies” include full-length antibodies, antigen-binding fragments of full-length antibodies, and molecules comprising antibody CDRs, VH regions, and / or VL regions. Examples of antibodies include, without limitation, monoclonal antibodies, recombinantly produced antibodies, monospecific antibodies, multispecific antibodies (including bispecific antibodies), human antibodies, humanized antibodies, chimeric antibodies, immunoglobulins, synthetic antibodies, tetrameric antibodies comprising two heavy chain and two light chain molecules, an antibody light chain monomer, an antibody heavy chain monomer, an antibody light chain dimer, an antibody heavy chain dimer, an antibody light chain-antibody heavy chain pair, intrabodies, heteroconjugate antibodies, antibody-drug conjugates, bispecific T cell engagers (BiTEs), chimeric antigen receptors, single domain antibodies, monovalent antibodies, single-chain antibodies or single-chain Fvs (scFv), camelized antibodies, affibodies, Fab fragments, F(ab’)2 fragments, disulfide-linked Fvs (sdFv), anti -idiotypic (anti-Id) antibodies (including, e.g., anti-anti -Id antibodies), and antigen-binding fragments of any of the above. Incertain embodiments, antibodies described herein refer to polyclonal antibody populations. Antibodies can be of any type (e.g., IgG, IgE, IgM, IgD, IgA, or IgY), any class (e.g., IgGi, IgG2, IgG3, IgG4, IgAi. or IgA2), or any subclass (e.g., IgG2a or IgG2b) of immunoglobulin molecule. In certain embodiments, antibodies described herein are IgG antibodies, or a class (e.g., human IgGi or IgG-t) or subclass thereof. In a specific embodiment, the antibody is a humanized monoclonal antibody. In another specific embodiment, the antibody is a human monoclonal antibody.

[0052] “Multispecific antibodies” are antibodies (e.g., bispecific antibodies) that specifically bind to two or more different antigens or two or more different regions of the same antigen. Multi specific antibodies include bispecific antibodies that contain two different antigen-binding sites (exclusive of the Fc region). Multispecific antibodies can include, for example, recombinantly produced antibodies, human antibodies, humanized antibodies, resurfaced antibodies, chimeric antibodies, immunoglobulins, synthetic antibodies, tetrameric antibodies comprising two heavy chain and two light chain molecules, an antibody light chain monomer, heteroconjugate antibodies, linked single-chain antibodies or linked-single-chain Fvs (scFv), camelized antibodies, affybodies, linked Fab fragments, F(ab’)2fragments, chemically-linked Fvs, and disulfide-linked Fvs (sdFv). Multi specific antibodies can be of any type (e.g., IgG, IgE, IgM, IgD, IgA, or IgY), any class (e.g., IgGi, lgG2, lgG3, lgG4, I Ai, or lgA2), or any subclass (e.g., lgG2a or lgG2b) of immunoglobulin molecule. In certain embodiments, multispecific antibodies described herein are IgG antibodies, or a class (e.g., human IgGi, IgG2, or IgG4) or subclass thereof.

[0053] As used herein, the term “CDR” or “complementarity determining region” means the noncontiguous antigen combining sites found within the variable regions of heavy and light chain polypeptides. These particular regions have been described by, for example, Kabat et al., J. Biol. Chem. 252, 6609-6616 (1977) and Kabat et al., Sequences of protein of immunological interest. (1991), by Chothia et al., J. Mol. Biol. 196: 901-917 (1987), and by MacCallum et al., J. Mol. Biol. 262: 732-745 (1996), all of which are herein incorporated by reference in their entireties, where the definitions include overlapping or subsets of amino acid residues when compared against each other. In certain embodiments, the term “CDR” is a CDR as defined by MacCallum et al., J. Mol. Biol. 262:732-745 (1996) and Martin A. “Protein Sequence and Structure Analysis of Antibody Variable Domains,” in Antibody Engineering, Kontermann and Diibel, eds., Chapter 31, pp. 422-439, Springer-Verlag, Berlin (2001). In certain embodiments, the term “CDR” is a CDR as defined by Kabat etal., J. Biol. Chem. 252, 6609-6616 (1977) and Kabat et al., Sequencesof protein of immunological interest. (1991). In certain embodiments, heavy chain CDRs and light chain CDRs of an antibody are defined using different conventions. In certain embodiments, heavy chain CDRs and / or light chain CDRs are defined by performing structural analysis of an antibody and identifying residues in the variable region(s) predicted to make contact with an epitope region of a target molecule (e.g., human LY6K). CDRH1, CDRH2, and CDRH3 denote the heavy chain CDRs, and CDRL1, CDRL2, and CDRL3 denote the light chain CDRs.

[0054] As used herein, the terms “variable region” and “variable domain” are used interchangeably and are common in the art. The variable region typically refers to a portion of an antibody, generally, a portion of a light or heavy chain, typically about the amino-terminal 110 to 120 amino acids or 110 to 125 amino acids in the mature heavy chain and about 90 to 115 amino acids in the mature light chain, which differ extensively in sequence among antibodies and are used in the binding and specificity of a particular antibody for its particular antigen. The variability in sequence is concentrated in those regions called complementarity determining regions (CDRs) while the more highly conserved regions in the variable region are called framework regions (FRs). Without wishing to be bound by any particular mechanism or theory, it is believed that the CDRs of the light and heavy chains are primarily responsible for the interaction and specificity of the antibody with antigen. In certain embodiments, the variable region is a human variable region. In certain embodiments, the variable region comprises rodent or murine CDRs and human framework regions (FRs). In certain embodiments, the variable region is a primate e.g., non-human primate) variable region. In certain embodiments, the variable region comprises rodent or murine CDRs and primate (e.g., non-human primate) framework regions (FRs).

[0055] As used herein, the terms “VH” and “VL” refer to antibody heavy and light chain variable regions, respectively, as described in Kabat et al., (1991) Sequences of Proteins of Immunological Interest (NUT Publication No. 91-3242, Bethesda), which is herein incorporated by reference in its entirety.

[0056] As used herein, the term “constant region” is common in the art. The constant region is an antibody portion, e.g., a carboxyl terminal portion of a light and / or heavy chain, which is not directly involved in binding of an antibody to antigen but which can exhibit various effector functions, such as interaction with an Fc receptor (e.g., Fc gamma receptor).

[0057] As used herein, the term “heavy chain” when used in reference to an antibody can refer to any distinct type, e.g., alpha (a), delta (5), epsilon (s), gamma (y), and mu (p), based on theamino acid sequence of the constant region, which give rise to IgA, IgD, IgE, IgG, and IgM classes of antibodies, respectively, including subclasses of IgG, e.., IgGi, IgG2, IgGs, and IgG4.

[0058] As used herein, the term “light chain” when used in reference to an antibody can refer to any distinct type, e.g., kappa (K) or lambda (X), based on the amino acid sequence of the constant region. Light chain amino acid sequences are well known in the art. In specific embodiments, the light chain is a human light chain.

[0059] As used herein, the term “specifically binds” refers to the specificity of a binding molecule (e.g., an antibody) for an antigen, as is understood by one skilled in the art. Binding molecules that specifically bind to an antigen typically bind to the antigen with an equilibrium dissociation constant (KD) of less than 1×10-6M, as measured by, e.g., ELISA assay, surface plasmon resonance, or other suitable assays known in the art. The skilled worker will appreciate that, in certain embodiments, a binding molecule can specifically bind to different antigens, e.g., different antigens that share a common epitope that is recognized by the binding molecule.

[0060] As used herein, the term “affinity” refers to the strength of the sum total of noncovalent interactions between a single binding site of a molecule (e.g., an antibody) and its binding partner (e.g., an antigen). Unless indicated otherwise, as used herein, “binding affinity” refers to intrinsic binding affinity which reflects a 1: 1 interaction between members of a binding pair (e.g., antibody and antigen). The affinity of a molecule X for its partner Y can generally be represented by the dissociation constant (KD). Affinity can be measured by common methods known in the art, including those described herein.

[0061] As used herein, the term “EU numbering system” refers to the EU numbering convention for the constant regions of an antibody, as described in Edelman, G. M. et al., Proc. Natl. Acad. USA, 63, 78-85 (1969) and Kabat et al, Sequences of Proteins of Immunological Interest, U. S. Dept. Health and Human Services, 5th edition, 1991, each of which is herein incorporated by reference in its entirety.

[0062] As used herein, the term “treat,” “treating,” and “treatment” refer to therapeutic or preventative measures described herein. The methods of “treatment” employ administration of an antibody to a subject having a disease or disorder, or predisposed to having such a disease or disorder, in order to prevent, cure, delay, reduce the severity of, or ameliorate one or more symptoms of the disease or disorder or recurring disease or disorder, or in order to prolong the survival of a subject beyond that expected in the absence of such treatment.

[0063] As used herein, the term “pharmaceutically active substance” refers to a molecule or moiety that is used to achieve a beneficial outcome in a subject. Beneficial outcomes include, but are not limited to, diagnosis, prognosis, treatment, cure and prevention (prophylaxis) of diseases and / or symptoms and / or health problems.

[0064] As used herein, the term “effective amount” in the context of the administration of a therapy to a subject refers to the amount of a therapy that achieves a desired prophylactic or therapeutic effect.

[0065] As used herein, the term “subject” includes any human or non-human animal. In certain embodiments, the subject is a human or non-human mammal. In certain embodiments, the subject is a human.

[0066] As used herein with respect to an antibody, polypeptide, or polynucleotide, the term “isolated” refers to an antibody, polypeptide, or polynucleotide that is separated from one or more contaminants (e.g., polypeptides, polynucleotides, lipids, or carbohydrates, etc.) which are present in a natural source (c.., in a mouse or a human) of the antibody, polypeptide, or polynucleotide. Moreover, an “isolated” antibody, polypeptide, or polynucleotide can be substantially free of other cellular material, or culture medium when produced by recombinant techniques, or substantially free of chemical precursors or other chemicals when chemically synthesized. For example, the language “substantially free” includes preparations of antibody, polypeptide, or polynucleotide having less than about 15%, 10%, 5%, 2%, 1%, 0.5%, or 0.1% (in particular less than about 10%) of other material, e.g., cellular material, culture medium, other nucleic acid molecules, chemical precursors, and / or other chemicals. All instances of “isolated antibodies” described herein are additionally contemplated as antibodies that may be, but need not be, isolated. All instances of “isolated polypeptides” described herein are additionally contemplated as polypeptides that may be, but need not be, isolated. All instances of “isolated polynucleotides” described herein are additionally contemplated as polynucleotides that may be, but need not be, isolated. All instances of “antibodies” described herein are additionally contemplated as antibodies that may be, but need not be, isolated. All instances of “polypeptides” described herein are additionally contemplated as polypeptides that may be, but need not be, isolated. All instances of “polynucleotides” described herein are additionally contemplated as polynucleotides that may be, but need not be, isolated.

[0067] The determination of “percent identity” between two sequences (e.g., amino acid sequences or nucleic acid sequences) can be accomplished using a mathematical algorithm. Aspecific, non-limiting example of a mathematical algorithm utilized for the comparison of two sequences is the algorithm of Karlin S & Altschul SF (1990) PNAS 87: 2264-2268, modified as in Karlin S & Altschul SF (1993) PNAS 90: 5873-5877, each of which is herein incorporated by reference in its entirety. Such an algorithm is incorporated into the NBLAST and XBLAST programs of Altschul SF et al., (1990) J Mol Biol 215: 403, which is herein incorporated by reference in its entirety. BLAST nucleotide searches can be performed with the NBLAST nucleotide program parameters set, e.g., for score=100, wordlength=12 to obtain nucleotide sequences homologous to a nucleic acid molecule described herein. BLAST protein searches can be performed with the XBLAST program parameters set, e.g., to score 50, wordlength=3 to obtain amino acid sequences homologous to a protein molecule described herein. To obtain gapped alignments for comparison purposes, Gapped BLAST can be utilized as described in Altschul SF et al., (1997) Nuc Acids Res 25: 3389-3402, which is herein incorporated by reference in its entirety. Alternatively, PSI BLAST can be used to perform an iterated search which detects distant relationships between molecules (Id.). When utilizing BLAST, Gapped BLAST, and PSI Blast programs, the default parameters of the respective programs (e.g., of XBLAST and NBLAST) can be used (see, e.g., National Center for Biotechnology Information (NCBI) on the worldwide web, ncbi.nlm.nih.gov). Another specific, non-limiting example of a mathematical algorithm utilized for the comparison of sequences is the algorithm of Myers and Miller, 1988, CABIOS 4: 11-17, which is herein incorporated by reference in its entirety. Such an algorithm is incorporated in the ALIGN program (version 2.0) which is part of the GCG sequence alignment software package. When utilizing the ALIGN program for comparing amino acid sequences, a PAM120 weight residue table, a gap length penalty of 12, and a gap penalty of 4 can be used.

[0068] The percent identity between two sequences can be determined using techniques similar to those described above, with or without allowing gaps. In calculating percent identity, typically only exact matches are counted.

[0069] As used herein, the term “linker” refers to a moiety that connects (e.g., covalently links) two or more elements of a compound. A linker may comprise one or more spacer moieties. As used herein, the term “spacer moiety” refers to a moiety that spaces (i.e., provides distance between) and covalently links together two (or more) parts of a linker. The linker may be part of, e.g., a linker-construct, a linker-conjugate, a linker-payload (e.g., linker-drug) or an antibody-drug conjugate, as defined below.

[0070] As used herein, the term “hydrophilic group” or “polar linker” refers to any molecular structure containing one or more polar functional groups that imparts improved polarity, and therefore improved aqueous solubility, to the molecule it is attached to. Hydrophilic groups may be selected from a carboxylic acid group, an alcohol group, an ether group, a polyethylene glycol group, an amino group, an ammonium group, a sulfonate group, a phosphate group, an acyl sulfamide group, and a carbamoyl sulfamide group. In addition to higher solubility, other effects of the hydrophilic group may include, without limitation, improved click conjugation efficiency, and, once incorporated into an antibody-drug conjugate: less aggregation, improved pharmacokinetics resulting in higher efficacy and in vivo tolerability.

[0071] As used herein, the term “salt thereof’ refers to a compound formed when an acidic proton, typically a proton of an acid, is replaced by a cation, such as a metal cation or an organic cation and the like. Where applicable, the salt is a pharmaceutically acceptable salt, although this is not required for salts that are not intended for administration to a subject. For example, in a salt of a compound, the compound may be protonated by an inorganic or organic acid to form a cation, with the conjugate base of the inorganic or organic acid as the anionic component of the salt. The term “pharmaceutically accepted” salt refers to a salt that is acceptable for administration to a subject, e.g., a mammal (z.c., salts with counterions having acceptable mammalian safety for a given dosage regime). Such salts may be derived from pharmaceutically acceptable inorganic or organic bases and from pharmaceutically acceptable inorganic or organic acids. As used herein, the term “pharmaceutically acceptable salt” refers to pharmaceutically acceptable salts of a compound, which salts are derived from a variety of organic and inorganic counter ions known in the art and include, for example, sodium, potassium, calcium, magnesium, ammonium, tetraalkylammonium, etc., and when the molecule contains a basic functionality, salts of organic or inorganic acids, such as hydrochloride, hydrobromide, formate, tartrate, besylate, mesylate, acetate, maleate, oxalate, etc.

[0072] As used herein, the terms “enediyne” or “enediyne antibiotic” or “enediyne-containing cytotoxin” refer to any cytotoxin characterized by the presence of a 3-ene-l, 5-diyne structural feature as part of a cyclic molecule as known in the art and include neocarzinostatin (NCS), C-1027, kedarcidin (KED), maduropeptin (MDP), N1999A2, the sporolides (SPO), the cyanosporasides (CYA and CYN), and the fijiolides, calicheamicins (CAL), the esperamicins (ESP), dynemicin (DYN), namenamicin, shishijimicin, and uncialamycin (UCM).

[0073] As used herein, the term “alkylaminosugar” refers to a tetrahydropyranyl moiety connected to an alcohol function via its 2-position, thereby forming an acetal function, and further substituted by (at least) one N-alkylamino group in position 3, 4 or 5. “N-alkylamino group” in this context refers to an amino group having one methyl, ethyl or 2-propyl group.

[0074] As used herein, the term “click probe” refers to a functional moiety that is capable of undergoing a click reaction, i.e., two compatible click probes mutually undergo a click reaction such that they are covalently linked in the product. Compatible probes for click reactions are known in the art, and include, without limitation, (cyclic) alkynes and azides. In the context of the present disclosure, click probe Q in a compound provided herein is capable of reacting with click probe F on a (modified) protein, such that upon the occurrence of a click reaction, a conjugate is formed wherein the protein is conjugated to the compound according to the present diclosure. Herein, F and Q are compatible click probes.

[0075] As used herein, “acylsulfamide moiety” refers to a sulfamide moiety (H2NSO2NH2) that is N-acylated or N-carbamoylated on one end of the molecule and N-alkylated (mono or bis) at the other end of the molecule. In the context of the present disclosure, this group is also referred to as “HS”.

[0076] As used herein, the term “glycoprotein” has its normal scientific meaning and refers to a protein comprising one or more monosaccharide or oligosaccharide chains (“glycans”) covalently bonded to the protein. A glycan may be attached to a hydroxyl group on the protein (O-linked-glycan), e.g., to the hydroxyl group of serine, threonine, tyrosine, hydroxylysine or hydroxyproline, or to an amide function on the protein (TV-gly coprotein), e.g., asparagine or arginine, or to a carbon on the protein (C-glycoprotein), e.g., tryptophan. A glycoprotein may comprise more than one glycan, may comprise a combination of one or more monosaccharide and one or more oligosaccharide glycans, and may comprise a combination of N-linked, O-linked and C-linked glycans. It is estimated that more than 50% of all proteins have some form of glycosylation and therefore qualify as glycoprotein. Examples of glycoproteins include PSMA (prostate-specific membrane antigen), CAL (candida antartica lipase), gp41, gpl20, EPO (erythropoietin), antifreeze protein and antibodies.

[0077] As used herein, the term “glycan” has its normal scientific meaning and refers to a monosaccharide or oligosaccharide chain that is linked to a protein. The term glycan thus refers to the carbohydrate-part of a glycoprotein. The glycan is attached to a protein via the C-l carbon ofone sugar, which may be without further substitution (monosaccharide) or may be further substituted at one or more of its hydroxyl groups (oligosaccharide). A naturally occurring glycan typically comprises 1 to about 10 saccharide moi eties. However, when a longer saccharide chain is linked to a protein, said saccharide chain is herein also considered a glycan. A glycan of a glycoprotein may be a monosaccharide. Typically, a monosaccharide glycan of a glycoprotein consists of a single N-acetylglucosamine (GlcNAc), glucose (Glc), mannose (Man) or fucose (Fuc) covalently attached to the protein. A glycan may also be an oligosaccharide. An oligosaccharide chain of a glycoprotein may be linear or branched. In an oligosaccharide, the sugar that is directly attached to the protein is called the core sugar. In an oligosaccharide, a sugar that is not directly attached to the protein and is attached to at least two other sugars is called an internal sugar. In an oligosaccharide, a sugar that is not directly attached to the protein but to a single other sugar, i.e., carrying no further sugar substituents at one or more of its other hydroxyl groups, is called the terminal sugar. For the avoidance of doubt, there may exist multiple terminal sugars in an oligosaccharide of a glycoprotein, but only one core sugar. A glycan may be an O-linked glycan, an N-linked glycan or a C-linked glycan. In an O-linked glycan, a monosaccharide or oligosaccharide glycan is bonded to an O-atom in an amino acid of the protein, typically via a hydroxyl group of serine (Ser) or threonine (Thr). In an N-linked glycan, a monosaccharide or oligosaccharide glycan is bonded to the protein via an N-atom in an amino acid of the protein, typically via an amide nitrogen in the side chain of asparagine (Asn) or arginine (Arg). In a C-linked glycan, a monosaccharide or oligosaccharide glycan is bonded to a C-atom in an amino acid of the protein, typically to a C-atom of tryptophan (Trp).Anti-LY6K Antibodies

[0078] In one aspect, the instant disclosure provides antibody-drug conjugates comprising an antibody that specifically bind to LY6K (e.g, human LY6K). The VH, VL, CDRH and CDRL amino acid sequences of exemplary anti-LY6K antibodies are set forth in Tables 1-4, respectively.Table 1. VH amino acid sequences of exemplary anti-LY6K antibodies.Table 2. VL amino acid sequences of exemplary anti-LY6K antibodies.Table 3. CDRH amino acid sequences of exemplary anti-LY6K antibodies.Table 4. CDRL amino acid sequences of exemplary anti-LY6K antibodies.

[0079] The individual CDRs of an antibody described herein can be determined according to any CDR numbering scheme known in the art.

[0080] In certain embodiments, one or more of the CDRs of an antibody described herein can be determined according to Kabat et al., J. Biol. Chem. 252, 6609-6616 (1977) and Kabat et al., Sequences of protein of immunological interest (1991), each of which is herein incorporated by reference in its entirety.

[0081] In certain embodiments, an antibody described herein comprises the CDRH1, CDRH2, and / or CDRH3 of a VH amino acid sequence set forth in any of SEQ ID NOs: 1-34 as determined by the Kabat numbering scheme. In certain embodiments, an antibody described herein comprises the CDRL1, CDRL2, and / or CDRL3 of a VL amino acid sequence set forth in any of SEQ ID NOs: 35-68 as determined by the Kabat numbering scheme.

[0082] In certain embodiments, one or more of the CDRs of an antibody described herein can be determined according to the Chothia numbering scheme, which refers to the location of immunoglobulin structural loops see, e.g., Chothia C & Lesk AM, (1987), J Mol Biol 196: 901 -917; Al-Lazikani B et al., (1997) J Mol Biol 273: 927-948; Chothia C et al., (1992) J Mol Biol 227: 799-817; Tramontano A et al., (1990) J Mol Biol 215(1): 175-82; and U. S. Patent No.7,709,226, all of which are herein incorporated by reference in their entireties).

[0083] In certain embodiments, an antibody described herein comprises the CDRH1, CDRH2, and / or CDRH3 of a VH amino acid sequence set forth in any of SEQ ID NOs: 1-34 as determined by the Chothia numbering system. In certain embodiments, an antibody described herein comprises the CDRL1, CDRL2, and / or CDRL3 of a VL amino acid sequence set forth in any of SEQ ID NOs: 35-68 as determined by the Chothia numbering system.

[0084] In certain embodiments, one or more of the CDRs of an antibody described herein can be determined according to MacCallum RM et al., (1996) J Mol Biol 262: 732-745, herein incorporated by reference in its entirety. See also, e.g., Martin A. “Protein Sequence and Structure Analysis of Antibody Variable Domains,” in Antibody Engineering, Kontermann and Diibel, eds., Chapter 31, pp. 422-439, Springer-Verlag, Berlin (2001), herein incorporated by reference in its entirety.

[0085] In certain embodiments, an antibody described herein comprises the CDRH1, CDRH2, and / or CDRH3 of a VH amino acid sequence set forth in any of SEQ ID NOs: 1-34 as determined by the MacCallum numbering system. In certain embodiments, an antibody described herein comprises the CDRL1, CDRL2, and / or CDRL3 of a VL amino acid sequence set forth in any of SEQ ID NOs: 35-68 as determined by the MacCallum numbering system.

[0086] In certain embodiments, the CDRs of an antibody described herein can be determined according to the IMGT numbering system as described in: Lefranc M-P, (1999) The Immunologist 7: 132-136; Lefranc M-P et al., (1999) Nucleic Acids Res 27: 209-212, each of which is herein incorporated by reference in its entirety; and Lefranc M-P et al., (2009) Nucleic Acids Res 37: D1006-D1012.

[0087] In certain embodiments, an antibody described herein comprises the CDRH1, CDRH2, and / or CDRH3 of a VH amino acid sequence set forth in any of SEQ ID NOs: 1-34 as determined by the IMGT numbering system. In certain embodiments, an antibody described herein comprises the CDRL1, CDRL2, and / or CDRL3 of a VL amino acid sequence set forth in any of SEQ ID NOs: 35-68 as determined by the IMGT numbering system.

[0088] In certain embodiments, the CDRs of an antibody described herein can be determined according to the AbM numbering scheme, which refers to AbM hypervariable regions, which represent a compromise between the Kabat CDRs and Chothia structural loops, and are used by Oxford Molecular’s AbM antibody modeling software (Oxford Molecular Group, Inc.), herein incorporated by reference in its entirety.

[0089] In certain embodiments, an antibody described herein comprises the CDRH1, CDRH2, and / or CDRH3 of a VH amino acid sequence set forth in any of SEQ ID NOs: 1-34 as determined by the AbM numbering scheme. In certain embodiments, an antibody described herein comprises the CDRL1, CDRL2, and / or CDRL3 of a VL amino acid sequence set forth in any of SEQ ID NOs: 35-68 as determined by the AbM numbering scheme.

[0090] In certain embodiments, the CDRs of an antibody described herein can be determined according to the AHo numbering system, as described in Honegger and Pliickthun, A., J. Mol. Biol. 309:657-670 (2001), herein incorporated by reference in its entirety.

[0091] In certain embodiments, an antibody described herein comprises the CDRH1, CDRH2, and / or CDRH3 of a VH amino acid sequence set forth in any of SEQ ID NOs: 1-34 as determined by the AHo numbering system. In certain embodiments, an antibody described herein comprises the CDRL1, CDRL2, and / or CDRL3 of a VL amino acid sequence set forth in any of SEQ ID NOs: 35-68 as determined by the AHo numbering system.

[0092] In certain embodiments, the individual CDRs of an antibody described herein are each independently determined according to one of the Kabat, Chothia, MacCallum, IMGT, AHo, or AbM numbering schemes, or by structural analysis of the multispecific molecule, wherein the structural analysis identifies residues in the variable region(s) predicted to make contact with an epitope region of LY6K.

[0093] In certain embodiments, the instant disclosure describes an antibody that specifically binds LY6K (e.g., human LY6K) comprising a VH comprising the CDRH1, CDRH2, and CDRH3 amino acid sequences of a VH amino acid sequence set forth in any one of SEQ ID NOs: 1-34, and a VL comprising the CDRL1, CDRL2, and CDRL3 amino acid sequences of a VL amino acid sequence set forth in any one of SEQ ID NOs: 35-68, wherein each CDR is independently determined according to one of the Kabat, Chothia, MacCallum, IMGT, AHo, or AbM numbering schemes, or by structural analysis of the multispecific molecule, wherein the structural analysis identifies residues in the variable region(s) predicted to make contact with an epitope region of LY6K (e.g., human LY6K).

[0094] In certain embodiments, the instant disclosure describes an antibody that specifically binds LY6K (e.g., human LY6K) comprising a VH comprising the CDRH1, CDRH2, and CDRH3 amino acid sequences of a VH amino acid sequence set forth in any one of SEQ ID NOs: 1-34, and a VL comprising the CDRL1, CDRL2, and CDRL3 amino acid sequences of a VL amino acid sequence set forth in any one of SEQ ID NOs: 35-68, wherein the VH amino acid sequence and the VL amino acid sequence are from the same antibody (i.e., as shown in Tables 1 and 2), and wherein each CDR is independently determined according to one of the Kabat, Chothia, MacCallum, IMGT, AHo, or AbM numbering schemes, or by structural analysis of themultispecific molecule, wherein the structural analysis identifies residues in the variable region(s) predicted to make contact with an epitope region of LY6K (e.g., human LY6K).

[0095] In certain embodiments, the instant disclosure describes an antibody that specifically binds to LY6K (e.g., human LY6K), wherein the antibody comprises the CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3 amino acid sequences of the VH and VL amino acid sequences set forth in SEQ ID NOs: 1 and 35; SEQ ID NOs: 2 and 36; SEQ ID NOs: 3 and 37; SEQ ID NOs: 4 and 38; SEQ ID NOs: 5 and 39; SEQ ID NOs: 6 and 40; SEQ ID NOs: 7 and 41; SEQ ID NOs: 8 and 42; SEQ ID NOs: 9 and 43; SEQ ID NOs: 10 and 44; SEQ ID NOs: 11 and 45; SEQ ID NOs: 12 and 46; SEQ ID NOs: 13 and 47; SEQ ID NOs: 14 and 48; SEQ ID NOs: 15 and 49; SEQ ID NOs: 16 and 50; SEQ ID NOs: 17 and 51; SEQ ID NOs: 18 and 52; SEQ ID NOs: 19 and 53; SEQ ID NOs: 20 and 54; SEQ ID NOs: 21 and 55; SEQ ID NOs: 22 and 56; SEQ ID NOs: 23 and 57; SEQ ID NOs: 24 and 58; SEQ ID NOs: 25 and 59; SEQ ID NOs: 26 and 60; SEQ ID NOs: 27 and 61; SEQ ID NOs: 28 and 62; SEQ ID NOs: 29 and 63; SEQ ID NOs: 30 and 64; SEQ ID NOs: 31 and 65; SEQ ID NOs: 32 and 66; SEQ ID NOs: 33 and 67; or SEQ ID NOs: 34 and 68, respectively.

[0096] In certain embodiments, the instant disclosure describes an antibody that specifically binds to LY6K (e.g., human LY6K), wherein the antibody comprises a VH comprising a CDRH1, CDRH2, and / or CDRH3 amino acid sequence set forth in Table 3. In certain embodiments, the antibody comprises a VH comprising a CDRH1 amino acid sequence set forth in any one of SEQ ID NOs: 69-89. In certain embodiments, the antibody comprises a VH comprising a CDRH2 amino acid sequence set forth in any one of SEQ ID NOs: 90-123. In certain embodiments, the antibody comprises a VH comprising a CDRH3 amino acid sequence set forth in any one of SEQ ID NOs: 124-157.

[0097] In certain embodiments, the instant disclosure describes an antibody that specifically binds to LY6K (e.g., human LY6K), wherein the antibody comprises a VH comprising the CDRH1, CDRH2, and CDRH3 amino acid sequences of any of the antibodies in Table 3. In certain embodiments, the antibody comprises a VH comprising the CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in SEQ ID NOs: 69, 90, and 124; SEQ ID NOs: 70, 91, and 125; SEQ ID NOs: 71, 92, and 126; SEQ ID NOs: 72, 93, and 127; SEQ ID NOs: 69, 94, and 124; SEQ ID NOs: 73, 95, and 128; SEQ ID NOs: 74, 96, and 129; SEQ ID NOs: 75, 97, and 130; SEQ ID NOs: 74, 98, and 131; SEQ ID NOs: 76, 99, and 132; SEQ ID NOs: 77, 100, and 133; SEQ ID NOs: 78,101, and 134; SEQ ID NOs: 79, 102, and 135; SEQ ID NOs: 74, 103, and 136; SEQ ID NOs: 80, 104, and 137; SEQ ID NOs: 74, 105, and 138; SEQ ID NOs: 74, 106, and 139; SEQ ID NOs: 81, 107, and 140; SEQ ID NOs: 81, 108, and 141; SEQ ID NOs: 82, 109, and 142; SEQ ID NOs: 83, 110, and 143; SEQ ID NOs: 84, 99, and 144; SEQ ID NOs: 85, 111, and 145; SEQ ID NOs: 81, 112, and 146; SEQ ID NOs: 86, 113, and 147; SEQ ID NOs: 87, 114, and 148; SEQ ID NOs: 74, 115, and 149; SEQ ID NOs: 74, 116, and 150; SEQ ID NOs: 74, 117, and 151; SEQ ID NOs: 88, 118, and 152; SEQ ID NOs: 89, 119, and 153; SEQ ID NOs: 81, 120, and 154; SEQ ID NOs: 74, 121, and 155; SEQ ID NOs: 85, 122, and 156; or SEQ ID NOs: 86, 123, and 157; respectively.

[0098] In certain embodiments, the instant disclosure describes an antibody that specifically binds to LY6K (e.g., human LY6K), wherein the antibody comprises a VL comprising a CDRL1, CDRL2, and / or CDRL3 amino acid sequence set forth in Table 4. In certain embodiments, the antibody comprises a VL comprising a CDRL1 amino acid sequence set forth in any one of SEQ ID NOs: 158-184. In certain embodiments, the antibody comprises a VL comprising a CDRL2 amino acid sequence set forth in any one of KIS, WAS, STT, AAS, and SEQ ID NOs: 185-197. In certain embodiments, the antibody comprises a VL comprising a CDRL3 amino acid sequence set forth in any one of SEQ ID NOs: 198-227.

[0099] In certain embodiments, the instant disclosure describes an antibody that specifically binds to LY6K (e.g., human LY6K), wherein the antibody comprises a VL comprising the CDRL1, CDRL2, and CDRL3 amino acid sequences of any of the antibodies in Table 4. In certain embodiments, the antibody comprises a VL comprising the CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in SEQ ID NO: 158, KIS, and SEQ ID NO: 198; SEQ ID NO: 159, WAS, and SEQ ID NO: 199; SEQ ID NO: 160, STT, and SEQ ID NO: 200; SEQ ID NO: 161, WAS, and SEQ ID NO: 201; SEQ ID NO: 162, AAS, and SEQ ID NO: 202; SEQ ID NOs: 163, 185, and 203; SEQ ID NOs: 164, 185, and 204; SEQ ID NOs: 165, 185, and 205; SEQ ID NOs: 166, 186, and 206; SEQ ID NOs: 167, 187, and 207; SEQ ID NOs: 168, 188, and 208; SEQ ID NOs: 169, 186, and 209; SEQ ID NOs: 170, 185, and 198; SEQ ID NOs: 169, 189, and 210; SEQ ID NOs: 170, 185, and 211; SEQ ID NOs: 171, 187, and 212; SEQ ID NOs: 172, 187, and 213; SEQ ID NOs: 173, 190, and 214; SEQ ID NOs: 174, 191, and 215; SEQ ID NOs: 175, 186, and 216; SEQ ID NOs: 176, 186, and 217; SEQ ID NOs: 170, 192, and 198; SEQ ID NOs: 171, 193, and 218; SEQ ID NOs: 177, 187, and 219; SEQ ID NOs: 178, 189, and 220; SEQ ID NOs: 179, 187, and 221; SEQ ID NOs: 165, 185, and 222; SEQ ID NOs: 180, 187, and 223; SEQ ID NOs: 181, 194,and 224; SEQ ID NOs: 182, 195, and 199; SEQ ID NOs: 172, 187, and 225; SEQ ID NOs: 183, 196, and 226; or SEQ ID NOs: 184, 197, and 227, respectively.

[0100] In certain embodiments, the instant disclosure describes an antibody that specifically binds to LY6K (e.g., human LY6K), wherein the antibody comprises a VH comprising a CDRH1, CDRH2, and / or CDRH3 amino acid sequence set forth in Table 3 and a VL comprising a CDRL1, CDRL2, and / or CDRL3 amino acid sequence set forth in Table 4, wherein 2, 3, 4, 5, or 6 of the CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3 amino acid sequences are from the same antibody (z.e., as shown in Tables 3 and 4). In certain embodiments, the antibody comprises a VH comprising CDRH1, CDRH2, and CDRH3 regions, and a VL comprising CDRL1, CDRL2, and CDRL3 regions, wherein the CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3 regions comprise the amino acid sequences set forth in SEQ ID NOs: 69, 90, 124, 158, KIS, and SEQ ID NO: 198; SEQ ID NOs: 70, 91, 125, 159, WAS, and SEQ ID NO: 199; SEQ ID NOs: 71, 92, 126, 160, STT, and SEQ ID NO: 200; SEQ ID NOs: 72, 93, 127, 161, WAS, and SEQ ID NO: 201; SEQ ID NOs: 69, 94, 124, 158, KIS, and SEQ ID NO: 198; SEQ ID NOs: 73, 95, 128, 162, AAS, and SEQ ID NO: 202; SEQ ID NOs: 74, 96, 129, 163, 185, and 203; SEQ ID NOs: 75, 97, 130, 164, 185, and 204; SEQ ID NOs: 74, 98, 131, 165, 185, and 205; SEQ ID NOs: 76, 99, 132, 166, 186, and 206; SEQ ID NOs: 77, 100, 133, 167, 187, and 207; SEQ ID NOs: 78, 101, 134, 168, 188, and 208; SEQ ID NOs: 79, 102, 135, 169, 186, and 209; SEQ ID NOs: 74, 103, 136, 170, 185, and 198; SEQ ID NOs: 80, 104, 137, 169, 189, and 210; SEQ ID NOs: 74, 105, 138, 170, 185, and 211; SEQ ID NOs: 74, 106, 139, 171, 187, and 212; SEQ ID NOs: 81, 107, 140, 172, 187, and 213; SEQ ID NOs: 81, 108, 141, 173, 190, and 214; SEQ ID NOs: 82, 109, 142, 174, 191, and 215; SEQ ID NOs: 83, 110, 143, 175, 186, and 216; SEQ ID NOs: 84, 99, 144, 176, 186, and 217; SEQ ID NOs: 85, 111, 145, 170, 192, and 198; SEQ ID NOs: 81, 112, 146, 171, 193, and 218; SEQ ID NOs: 86, 113, 147, 177, 187, and 219; SEQ ID NOs: 87, 114, 148, 178, 189, and 220; SEQ ID NOs: 74, 115, 149, 179, 187, and 221; SEQ ID NOs: 74, 116, 150, 165, 185, and 222; SEQ ID NOs: 74, 117, 151, 180, 187, and 223; SEQ ID NOs: 88, 118, 152, 181, 194, and 224; SEQ ID NOs: 89, 119, 153, 182, 195, and 199; SEQ ID NOs: 81, 120, 154, 172, 187, and 225; SEQ ID NOs: 74, 121, 155, 165, 185, and 222; SEQ ID NOs: 85, 122, 156, 183, 196, and 226; or SEQ ID NOs: 86, 123, 157, 184, 197, and 227, respectively.

[0101] In certain embodiments, the instant disclosure describes an antibody that specifically binds to LY6K (e.., human LY6K) comprising a VH comprising an amino acid sequence that isat least 75%, 80%, 85%, 90%, 95%, or 100% (e.g., at least 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, or 99%) identical to the amino acid sequence set forth in any one of SEQ ID NOs: 1-34. In certain embodiments, the instant disclosure describes an antibody that specifically binds to LY6K (e.g., human LY6K), comprising a VH comprising an amino acid sequence set forth in any one of SEQ ID NOs: 1-34. In certain embodiments, the amino acid sequence of the VH consists of the amino acid sequence set forth in any one of SEQ ID NOs: 1-34.

[0102] In certain embodiments, the instant disclosure describes an antibody that specifically binds to LY6K (e.g., human LY6K), comprising a VL comprising an amino acid sequence that is at least 75%, 80%, 85%, 90%, 95%, or 100% (e.g, at least 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, or 99%) identical to the amino acid sequence set forth in any one of SEQ ID NOs: 35-68. In certain embodiments, the instant disclosure describes an antibody that specifically binds to LY6K (e.g., human LY6K), comprising a VL comprising an amino acid sequence set forth in any one of SEQ ID NOs: 35-68. In certain embodiments, the amino acid sequence of the VL consists of the amino acid sequence set forth in any one of SEQ ID NOs: 35-68.

[0103] In certain embodiments, the instant disclosure describes an antibody that specifically binds to LY6K (e.g., human LY6K), comprising a VH comprising an amino acid sequence that is at least 75%, 80%, 85%, 90%, 95%, or 100% (e.g, at least 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98 or 99%) identical to the amino acid sequence set forth in any one of SEQ ID NOs: 1-34, and a VL comprising an amino acid sequence that is at least 75%, 80%, 85%, 90%, 95%, or 100% (e.g, at least 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, or 99%) identical to the amino acid sequence set forth in any one of SEQ ID NOs: 35-68. In certain embodiments, the instant disclosure describes an antibody that specifically binds to LY6K (e.g, human LY6K), comprising a VH comprising an amino acid sequence of any one of SEQ ID NOs: 1-34, and a VL comprising an amino acid sequence of any one of SEQ ID NOs: 35-68. In certain embodiments, the amino acid sequence of the VH consists of the amino acid sequence set forth in any one of SEQ ID NOs: 1-34, and the amino acid sequence of the VL consists of the amino acid sequence set forth in any one of SEQ ID NOs: 35-68.

[0104] In certain embodiments, the instant disclosure describes an antibody that specifically binds to LY6K (e.g., human LY6K), comprising a VH comprising an amino acid sequence that is at least 75%, 80%, 85%, 90%, 95%, or 100% (e.g., at least 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98 or 99%) identical to an amino acid sequence set forth in any one of SEQ ID NOs: 1-34,and a VL comprising an amino acid sequence that is at least 75%, 80%, 85%, 90%, 95%, or 100% (e.g., at least 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, or 99%) identical to an amino acid sequence set forth in any one of SEQ ID NOs: 35-68, wherein the amino acid sequences set forth in any one of SEQ ID NOs: 1-34 and any one of SEQ ID NOs: 35-68 are from the same antibody (z.e., as shown in Tables 1 and 2). In certain embodiments, the instant disclosure describes an antibody that specifically binds to LY6K (e.g., human LY6K), comprising a VH comprising an amino acid sequence of any one of SEQ ID NOs: 1-34, and a VL comprising an amino acid sequence of any one of SEQ ID NOs: 35-68, wherein the amino acid sequences set forth in any one of SEQ ID NOs: 1-34 and any one of SEQ ID NOs: 35-68 are from the same antibody (i.e., as shown in Tables 1 and 2). In certain embodiments, the amino acid sequence of the VH consists of the amino acid sequence set forth in any one of SEQ ID NOs: 1-34, and the amino acid sequence of the VL consists of the amino acid sequence set forth in any one of SEQ ID NOs: 35-68, wherein the amino acid sequences set forth in any one of SEQ ID NOs: 1-34 and any one of SEQ ID NOs: 35-68 are from the same antibody i.e., as shown in Tables 1 and 2).

[0105] In certain embodiments, the instant disclosure describes an antibody that specifically binds to LY6K (e.g., human LY6K), wherein the antibody comprises a VH and VL comprising the amino acid sequences set forth in SEQ ID NOs: 1 and 35; SEQ ID NOs: 2 and 36; SEQ ID NOs: 3 and 37; SEQ ID NOs: 4 and 38; SEQ ID NOs: 5 and 39; SEQ ID NOs: 6 and 40; SEQ ID NOs: 7 and 41; SEQ ID NOs: 8 and 42; SEQ ID NOs: 9 and 43; SEQ ID NOs: 10 and 44; SEQ ID NOs: 11 and 45; SEQ ID NOs: 12 and 46; SEQ ID NOs: 13 and 47; SEQ ID NOs: 14 and 48; SEQ ID NOs: 15 and 49; SEQ ID NOs: 16 and 50; SEQ ID NOs: 17 and 51; SEQ ID NOs: 18 and 52; SEQ ID NOs: 19 and 53; SEQ ID NOs: 20 and 54; SEQ ID NOs: 21 and 55; SEQ ID NOs: 22 and 56; SEQ ID NOs: 23 and 57; SEQ ID NOs: 24 and 58; SEQ ID NOs: 25 and 59; SEQ ID NOs: 26 and 60; SEQ ID NOs: 27 and 61; SEQ ID NOs: 28 and 62; SEQ ID NOs: 29 and 63; SEQ ID NOs: 30 and 64; SEQ ID NOs: 31 and 65; SEQ ID NOs: 32 and 66; SEQ ID NOs: 33 and 67; or SEQ ID NOs: 34 and 68, respectively.

[0106] In certain embodiments, the instant disclosure describes an antibody that specifically binds to LY6K (e.g., human LY6K), wherein the antibody comprises a VH and VL consisting of the amino acid sequences set forth in SEQ ID NOs: 1 and 35; SEQ ID NOs: 2 and 36; SEQ ID NOs: 3 and 37; SEQ ID NOs: 4 and 38; SEQ ID NOs: 5 and 39; SEQ ID NOs: 6 and 40; SEQ ID NOs: 7 and 41; SEQ ID NOs: 8 and 42; SEQ ID NOs: 9 and 43; SEQ ID NOs: 10 and 44; SEQ IDNOs: 11 and 45; SEQ ID NOs: 12 and 46; SEQ ID NOs: 13 and 47; SEQ ID NOs: 14 and 48; SEQ ID NOs: 15 and 49; SEQ ID NOs: 16 and 50; SEQ ID NOs: 17 and 51; SEQ ID NOs: 18 and 52; SEQ ID NOs: 19 and 53; SEQ ID NOs: 20 and 54; SEQ ID NOs: 21 and 55; SEQ ID NOs: 22 and 56; SEQ ID NOs: 23 and 57; SEQ ID NOs: 24 and 58; SEQ ID NOs: 25 and 59; SEQ ID NOs: 26 and 60; SEQ ID NOs: 27 and 61; SEQ ID NOs: 28 and 62; SEQ ID NOs: 29 and 63; SEQ ID NOs: 30 and 64; SEQ ID NOs: 31 and 65; SEQ ID NOs: 32 and 66; SEQ ID NOs: 33 and 67; or SEQ ID NOs: 34 and 68, respectively.

[0107] In certain embodiments, the instant disclosure describes an antibody that crosscompetes for binding to LY6K (e.g., human LY6K) with any of the antibodies described above. In certain embodiments, the instant disclosure describes an antibody that binds to the same or an overlapping epitope of LY6K (e.g., an epitope of human LY6K) as an antibody described above.

[0108] In certain embodiments, the epitope of an antibody can be determined by, e.g., NMR spectroscopy, surface plasmon resonance (BIAcore®), X-ray diffraction crystallography studies, ELISA assays, hydrogen / deuterium exchange coupled with mass spectrometry (e.g., liquid chromatography electrospray mass spectrometry), array-based oligo-peptide scanning assays, and / or mutagenesis mapping (e.g., site-directed mutagenesis mapping). For X-ray crystallography, crystallization may be accomplished using any of the known methods in the art (e.g., Giege R et al., (1994) Acta Crystallogr D Biol Crystallogr 50(Pt 4): 339-350; McPherson A (1990) Eur J Biochem 189: 1-23; Chayen NE (1997) Structure 5: 1269-1274; McPherson A (1976) J Biol Chem 251: 6300-6303, all of which are herein incorporated by reference in their entireties). Antibody: antigen crystals may be studied using well known X-ray diffraction techniques and may be refined using computer software such as X-PLOR (Yale University, 1992, distributed by Molecular Simulations, Inc.; see, e.g., Meth Enzymol (1985) volumes 114 & 115, eds. Wyckoff HW et al, U. S. Patent Application No. 2004 / 0014194), and BUSTER (Bricogne G (1993) Acta Crystallogr D Biol Crystallogr 49(Pt 1): 37-60; Bricogne G (1997) Meth Enzymol 276A: 361-423, ed Carter CW; Roversi P etal., (2000) Acta Crystallogr D Biol Crystallogr 56(Pt 10): 1316-1323, all of which are herein incorporated by reference in their entireties). Mutagenesis mapping studies may be accomplished using any method known to one of skill in the art. See, e.g., Champe M et al., (1995) supra and Cunningham BC & Wells JA (1989) supra for a description of mutagenesis techniques, including alanine scanning mutagenesis techniques. In a specific embodiment, the epitope of an antibody is determined using alanine scanning mutagenesis studies. In addition, orantibodies that recognize and bind to the same or overlapping epitopes of LY6K (e.., human LY6K) can be identified using routine techniques such as an immunoassay, for example, by showing the ability of one antibody to block the binding of another antibody to a target antigen, i.e., a competitive binding assay. Competition binding assays also can be used to determine whether two antibodies have similar binding specificity for an epitope. Competitive binding can be determined in an assay in which the immunoglobulin under test inhibits specific binding of a reference antibody to a common antigen, such as LY6K (e.g., human LY6K). Numerous types of competitive binding assays are known, for example: solid phase direct or indirect radioimmunoassay (RIA), solid phase direct or indirect enzyme immunoassay (EIA), sandwich competition assay (see Stahli C et al., (1983) Methods Enzymol 9: 242-253); solid phase direct biotin-avidin EIA (see Kirkland TN et al., (1986) J Immunol 137: 3614-9); solid phase direct labeled assay, solid phase direct labeled sandwich assay (see Harlow E & Lane D, (1988) Antibodies: A Laboratory Manual, Cold Spring Harbor Press); solid phase direct label RIA using 1-125 label (see Morel GA et al., (1988) Mol Immunol 25(1): 7-15); solid phase direct biotinavidin EIA (see Cheung RC et al., (1990) Virology 176: 546-52); and direct labeled RIA (see Moldenhauer G et al., (1990) Scand J Immunol 32: 77-82), all of which are herein incorporated by reference in their entireties. Typically, such an assay involves the use of purified antigen (e.g., LY6K, such as human LY6K) bound to a solid surface or cells bearing either of these, an unlabeled test immunoglobulin and a labeled reference immunoglobulin. Competitive inhibition can be measured by determining the amount of label bound to the solid surface or cells in the presence of the test immunoglobulin. Usually, the test immunoglobulin is present in excess. Usually, when a competing antibody is present in excess, it will inhibit specific binding of a reference or antibody to a common antigen by at least 50-55%, 55-60%, 60-65%, 65-70%, 70-75%, or more. A competition binding assay can be configured in a large number of different formats using either labeled antigen or labeled antibody. In a common version of this assay, the antigen is immobilized on a 96-well plate. The ability of unlabeled antibodies to block the binding of labeled antibodies to the antigen is then measured using radioactive or enzyme labels. For further details see, e.g., W agener C et al., (1983) J Immunol 130: 2308-2315; Wagener C etal., (1984) J Immunol Methods 68: 269-274; Kuroki M etal., (1990) Cancer Res 50: 4872-4879; Kuroki Me / al., (1992) Immunol Invest 21: 523-538; Kuroki Metal., (1992) Hybridoma 11: 391-407 and Antibodies: A LaboratoryManual, Ed Harlow E & Lane D editors supra, pp. 386-389, all of which are herein incorporated by reference in their entireties.

[0109] The anti-LY6K antigen-binding molecules of the present disclosure can be linked to or co-expressed with another functional molecule, e.g., another peptide or protein. For example, an antibody or fragment thereof can be functionally linked (e.g., by chemical coupling, genetic fusion, noncovalent association, or otherwise) to one or more other molecular entities, such as another antibody or antibody fragment to produce a bispecific or a multispecific antibody (e.g., a bispecific T cell engager [BiTE] or a dual-affinity re-targeting antibody [DART]) with a second or additional binding specificity. In some embodiments, the bispecific or multispecific antibody has binding specificity for a molecule on an effector cell (e.g, CD3, CD16, CD137). In some embodiments, an antibody described herein is a multispecific antibody. In some embodiments, a multispecific antibody described herein comprises a CD3 -binding region. In some embodiments, the CD3-binding region comprises a sequence from an antibody known to bind CD3 (e.g., OKT3, UTCH1, L2K, TR66, etc.). In some embodiments, the CD3-binding region comprises a sequence from an L2K anti-CD3 antibody. In some embodiments, the CD3-binding region comprises a sequence from a de-immunized L2K anti-CD3 antibody. In some embodiments, the CD3 -binding region comprises a single-chain fragment variable (scFv) that specifically binds to CD3. In some embodiments, a multispecific antibody described herein specifically binds to human CD3.

[0110] In certain embodiments, an antibody described herein is conjugated to a pharmaceutically active substance (e.g., as described in the Antibody-drug Conjugates section below). Pharmaceutically active substances include, but are not limited to, cytotoxic agents, cytostatic agents, toxins, radionuclides (e.g., radioisotopes), polypeptides, polynucleoctides, detectable labels, and combinations thereof. In certain embodiments, pharmaceutically active substance is a cytotoxic agent. In certain embodiments, the cytotoxic agent is able to induce death or destruction of a cell in contact therewith. In certain embodiments, the pharmaceutically active substance is a cytostatic agent. In certain embodiments, the cytostatic agent is able to prevent or substantially reduce proliferation and / or inhibits the activity or function of a cell in contact therewith. In certain embodiments, the cytotoxic agent or cytostatic agent is a chemotherapeutic agent.

[0111] In certain embodiments, the pharmaceutically active substance is a radionuclide. Suitable radionuclides include, but are not limited to, beta emitters, auger emitters, convertedelectron emitters, alpha emitters, and low photon energy emitters. In certain embodiments, the radionuclide is selected from3H,14C,18F32P,35S,36C1,45Ca,51Cr,57Co,38Co,59Fe,64Cu,67Cu,67Ga,68Ga,76As,77As,89Sr,89Zr,90Y, "Tc, "mTc,105Rh,luIn,114mIn,117Lu,121I,1231,124I,1251,131I,149Tb,153Sm,161Tb,166HO,177LU,198AU,2O1T1,211At,212Pb,213Bi,225Ac,186Re,188Re,212Bi,213Bi,221At,223Ac,223Ra,225Ac,255Fm, and combinations thereof.

[0112] In certain embodiments, the pharmaceutically active substance is a detectable label. In certain embodiments, the detectable label comprises a fluorescent moiety, a click chemistry handle, or a combination thereof.

[0113] In certain embodiments, the pharmaceutically active substance is a drug. Suitable drugs include, but are not limited to, anti-cancer agents, anti-inflammatory agents, and anti-infective (e.g., anti-fungal, antibacterial, anti-parasitic, antiviral) agents. Suitable anti-cancer agents include, but are not limited to, alkylating agents, antimetabolites, spindle poison plant alkaloids, cytotoxic / antitumor antibiotics, topoisomerase inhibitors, photosensitizers, kinase inhibitors, anti-hormonal agents, aromatase inhibitors, anti-androgens, protein kinase inhibitors, lipid kinase inhibitors, antisense oligonucleotides (e.g., those which inhibit expression of genes in signaling pathways implicated in aberrant cell proliferation), ribozymes, (e.g., VEGF expression inhibitors and HER2 expression inhibitors), vaccines e.g., gene therapy vaccines), topoisomerase 1 inhibitors, anti -angiogenic agents, pharmaceutically acceptable salts, acids, solvates and derivatives of any of the above, and any combination thereof.

[0114] In certain embodiments, the pharmaceutically active substance is a toxin. Suitable toxins include, but are not limited to, proteinaceous toxins (e.., bacterial-derived toxins, and plant-derived toxins), toxins targeting tubulin filaments, toxins targeting DNA, toxins targeting RNA. Examples of proteinaceous toxins include saporin, dianthin, ricin, modeccin, abrin, volkensin, viscumin, shiga toxin, shiga-like toxin, pseudomonas exotoxin (PE, also known as exotoxin A), diphtheria toxin (DT), and cholera toxin. Examples of toxins targeting tubulin filaments include maytansinoids e.g., DM1 and DM4), auristatins (e.g., Monomethyl auristatin E (MMAE) and Monomethyl auristatin F (MMAF)), toxoids, tubulysins, cryptophycins, rhizoxin. Examples of DNA-targeting toxins include calicheamicins: N-Acetyl- y-calicheamicin, CC-1065 analogs, duocarmycins, doxorubicin, methotrexate, benzodiazepines, camptothecin analogues, and anthracyclines. Examples of RNA-targeting toxins are amanitins, spliceostatins, and thailanstatins.

[0115] In certain embodiments, the pharmaceutically active substance is a polypeptide. Suitable polypeptides include, but are not limited to, Cas9; toxins (e.g., saporin, dianthin, gelonin, (de)bouganin, agrostin, ricin (toxin A chain); pokeweed antiviral protein, apoptin, diphtheria toxin, pseudomonas exotoxin); metabolic enzymes (e.g., argininosuccinate lyase, argininosuccinate synthetase); enzymes of the coagulation cascade; repairing enzymes; enzymes for cell signaling; cell cycle regulation factors; gene regulating factors (e.g., ranscription factors such as NF -KB or gene repressors such as methionine repressor).

[0116] In certain embodiments, the pharmaceutically active substance is a polynucleotide. In certain embodiments, the polynucleotide comprises coding information. In certain embodiments, the polynucleotide is a gene or an open reading frame encoding a protein. In certain embodiments, the polynucleotide comprises regulatory information. In certain embodiments, the polynucleotide is a promoter, a regulatory element binding region, or a sequence encoding a micro RNA. Suitable polynucleotides include natural and artificial nucleic acids. Artificial nucleic acids include, but are not limited to, peptide nucleic acids (PNA), Morpholines and locked nucleic acids (LNA), glycol nucleic acids (GNA), and threose nucleic acids (TNA). Each of these is distinguished from naturally occurring DNA or RNA by changes to the backbone of the molecule. Suitable polynucleotides include, but are not limited to, a vector; a gene (e.g., a cell suicide-inducing transgene); single stranded DNA; linear double stranded DNA; circular double stranded DNA (e.g., a plasmid); mini-circle DNA; a DNA aptamer; single stranded RNA; linear double stranded RNA; mRNA; tRNA; rRNA; short interfering RNA (siRNA); microRNA (miRNA); antisense RNA; anti-sense oligonucleotides; peptide nucleic acid (PNA); phosphoramidate morpholino oligomer (PMO); locked nucleic acid (LNA); bridged nucleic acid (BNA); 2’-deoxy-2’-fluoroarabino nucleic acid (FANA); 2’-O-methoxyethyl-RNA (MOE); 2'-0,4'-aminoethylene bridged nucleic acid; 3 ’-fluoro hexitol nucleic acid (FHNA); an RNA aptamer; and combinations thereof.

[0117] Any immunoglobulin (Ig) constant region can be used in the antibodies described herein. In certain embodiments, the Ig region is a human IgG, IgE, IgM, IgD, IgA, or IgY immunoglobulin molecule, any class (e.g., IgGi, IgG2, IgGs, IgG4, IgAi, and IgA2), or any subclass (e.g., IgG2a and IgG2b) of immunoglobulin molecule.

[0118] In certain embodiments, the instant disclosure describes an antibody that specifically binds to LY6K (e.g., human LY6K), the antibody comprising a heavy chain constant region, optionally selected from the group consisting of human IgGi, IgG2, IgGs, IgG4, IgAi, and IgA2.

[0119] In certain embodiments, the instant disclosure describes an antibody that specifically binds to LY6K (e.g., human LY6K), the antibody comprising a heavy chain constant region that is a variant of a wild-type heavy chain constant region, wherein the variant heavy chain constant region binds to an FcyR with lower affinity than the wild-type heavy chain constant region binds to the FcyR.

[0120] In certain embodiments, the instant disclosure describes an antibody that specifically binds to LY6K (e.g., human LY6K), the antibody comprising a heavy chain constant region comprising an amino acid sequence shown in Table 5. In certain embodiments, the heavy chain constant region comprises the amino acid sequence of SEQ ID NO: 228 or 229. In certain embodiments, the heavy chain constant region consists of the amino acid sequence of SEQ ID NO: 228 or 229.Table 5. Heavy chain constant region amino acid sequences of exemplary anti-LY6K antibodies.

[0121] In certain embodiments, the instant disclosure describes an antibody that specifically binds to LY6K (e.g., human LY6K), the antibody comprising a light chain constant region comprising an amino acid sequence shown in Table 6. In certain embodiments, the heavy chain constant region comprises the amino acid sequence of SEQ ID NO: 230 or 231. In certainembodiments, the heavy chain constant region consists of the amino acid sequence of SEQ ID NO: 230 or 231.Table 6. Light chain constant region amino acid sequences of exemplary anti-LY6K antibodies.

[0122] In certain embodiments, one, two, or more mutations (e.g., amino acid substitutions) are introduced into an Fc region (e.g., a CH2 domain (residues 231-340 of human IgGi)) and / or a CH3 domain (residues 341-447 of human IgGi, numbered according to the EU numbering system) and / or a hinge region (residues 216-230, numbered according to the EU numbering system) of an antibody described herein, to alter one or more functional properties of the antibody, such as serum half-life, complement fixation, Fc receptor binding, and / or antigen-dependent cellular cytotoxicity.

[0123] In certain embodiments, one, two, or more mutations (e.g., amino acid substitutions) are introduced into the hinge region of an antibody described herein, such that the number of cysteine residues in the hinge region is altered (e.g., increased or decreased) as described in, e.g., U. S. Patent No. 5,677,425, herein incorporated by reference in its entirety. The number of cysteine residues in the hinge region may be altered to, e.g., facilitate assembly of the light and heavy chains, or to alter (e.g., increase or decrease) the stability of the antibody.

[0124] In a specific embodiment, one, two, or more amino acid mutations (e.g., substitutions, insertions, or deletions) are introduced into an IgG constant region, or FcRn-binding fragment thereof (preferably an Fc or hinge-Fc fragment) to alter (e.g., decrease or increase) half-life of the antibody in vivo. See, e.g., International Publication Nos. WO 02 / 060919; WO 98 / 23289; and WO 97 / 34631; and U. S. Patent Nos. 5,869,046, 6,121,022, 6,277,375, and 6,165,745, all of which are herein incorporated by reference in their entireties, for examples of mutations that will alter (e.g., decrease or increase) the half-life of an antibody in vivo. In certain embodiments, one, two or more amino acid mutations (e.g., substitutions, insertions, or deletions) are introduced into an IgGconstant region, or FcRn-binding fragment thereof (preferably an Fc or hinge-Fc fragment) to decrease the half-life of the antibody in vivo. In other embodiments, one, two or more amino acid mutations (e.g., substitutions, insertions, or deletions) are introduced into an IgG constant region, or FcRn-binding fragment thereof (preferably an Fc or hinge-Fc fragment) to increase the half-life of the antibody in vivo. In a specific embodiment, the antibodies may have one or more amino acid mutations (e.g., substitutions) in the second constant (CH2) domain (residues 231-340 of human IgGi) and / or the third constant (CH3) domain (residues 341-447 of human IgGi), numbered according to the EU numbering system. In a specific embodiment, the constant region of the IgGi of antibody described herein comprises a methionine (M) to tyrosine (Y) substitution in position 252, a serine (S) to threonine (T) substitution in position 254, and a threonine (T) to glutamic acid (E) substitution in position 256, numbered according to the EU numbering system. See U. S. Patent No. 7,658,921, which is herein incorporated by reference in its entirety. This type of mutant IgG, referred to as “YTE mutant” has been shown to display fourfold increased half-life as compared to wild-type versions of the same antibody (see Dall’Acqua WF el al., (2006) J Biol Chem 281: 23514-24, which is herein incorporated by reference in its entirety). In certain embodiments, an antibody comprises an IgG constant region comprising one, two, three, or more amino acid substitutions of amino acid residues at positions 251-257, 285-290, 308-314, 385-389, and 428-436, numbered according to the EU numbering system.

[0125] In certain embodiments, one, two, or more mutations (e.g., amino acid substitutions) are introduced into an Fc region (e.g, a CH2 domain (residues 231-340 of human IgGi)) and / or a CH3 domain (residues 341-447 of human IgGi, numbered according to the EU numbering system) and / or a hinge region (residues 216-230, numbered according to the EU numbering system) of an antibody described herein, to increase or decrease the affinity of the antibody for an Fc receptor e.g., an activated Fc receptor) on the surface of an effector cell. Mutations in the Fc region of an antibody that decrease or increase the affinity of an antibody for an Fc receptor and techniques for introducing such mutations into the Fc receptor or fragment thereof are known to one of skill in the art. Examples of mutations in the Fc receptor of an antibody that can be made to alter the affinity of the antibody for an Fc receptor are described in, e.g., Smith P et al., (2012) PNAS 109: 6181-6186, U. S. Patent No. 6,737,056, and International Publication Nos. WO 02 / 060919; WO 98 / 23289; and WO 97 / 34631, all of which are herein incorporated by reference in their entireties.

[0126] In certain embodiments, the antibody comprises a heavy chain constant region that is a variant of a wild-type heavy chain constant region, wherein the variant heavy chain constant region binds to FcyRIIB with higher affinity than the wild-type heavy chain constant region binds to FcyRIIB. In certain embodiments, the variant heavy chain constant region is a variant human heavy chain constant region, e.g., a variant human IgGi, a variant human IgG2, or a variant human IgG4 heavy chain constant region. In certain embodiments, the variant human IgG heavy chain constant region comprises one or more of the following amino acid mutations, according to the EU numbering system: G236D, P238D, S239D, S267E, L328F, and L328E. In certain embodiments, the variant human IgG heavy chain constant region comprises a set of amino acid mutations selected from the group consisting of: S267E and L328F; P238D and L328E; P238D and one or more substitutions selected from the group consisting of E233D, G237D, H268D, P271G, and A330R; P238D, E233D, G237D, H268D, P271G, and A330R; G236D and S267E; S239D and S267E; V262E, S267E, and L328F; and V264E, S267E, and L328F, according to the EU numbering system. In certain embodiments, the FcyRIIB is expressed on a cell selected from the group consisting of macrophages, monocytes, B cells, dendritic cells, endothelial cells, and activated T cells.[00127J In a further embodiment, one, two, or more amino acid substitutions are introduced into an IgG constant region Fc region to alter the effector function(s) of the antibody. For example, one or more amino acids selected from amino acid residues 234, 235, 236, 237, 239, 243, 267, 292, 297, 300, 318, 320, 322, 328, 330, 332, and 396, numbered according to the EU numbering system, can be replaced with a different amino acid residue such that the antibody has an altered affinity for an effector ligand but retains the antigen-binding ability of the parent antibody. The effector ligand to which affinity is altered can be, for example, an Fc receptor or the Cl component of complement. This approach is described in further detail in U. S. Patent Nos. 5,624,821 and 5,648,260, each of which is herein incorporated by reference in its entirety. In certain embodiments, the deletion or inactivation (through point mutations or other means) of a constant region domain may reduce Fc receptor binding of the circulating antibody thereby increasing tumor localization. See, e.g., U. S. Patent Nos. 5,585,097 and 8,591,886, each of which is herein incorporated by reference in its entirety, for a description of mutations that delete or inactivate the constant region and thereby increase tumor localization. In certain embodiments, one or more amino acid substitutions may be introduced into the Fc region of an antibody described herein toremove potential glycosylation sites on the Fc region, which may reduce Fc receptor binding (see, e.g., Shields RL et al., (2001) J Biol Chem 276: 6591-604, which is herein incorporated by reference in its entirety). In various embodiments, one or more of the following mutations in the constant region of an antibody described herein may be made: an N297A substitution; an N297Q substitution; an L234A substitution; an L234F substitution; an L235A substitution; an L235F substitution; an L235V substitution; an L237A substitution; an S239D substitution; an E233P substitution; anL234V substitution; a C236 deletion; aP238A substitution; anF243L substitution; a D265A substitution; an S267E substitution; an L328F substitution; an R292P substitution; a Y300L substitution; an A327Q substitution; a P329A substitution; an A330L substitution; an I332E substitution; or a P396L substitution, numbered according to the EU numbering system.

[0128] In certain embodiments, a mutation selected from the group consisting of D265A, P329A, and a combination thereof, numbered according to the EU numbering system, may be made in the constant region of an antibody described herein. In certain embodiments, a mutation selected from the group consisting of L235A, L237A, and a combination thereof, numbered according to the EU numbering system, may be made in the constant region of an antibody described herein. In certain embodiments, a mutation selected from the group consisting of S267E, L328F, and a combination thereof, numbered according to the EU numbering system, may be made in the constant region of an antibody described herein. In certain embodiments, a mutation selected from the group consisting of S239D, I332E, optionally A330L, and a combination thereof, numbered according to the EU numbering system, may be made in the constant region of an antibody described herein. In certain embodiments, a mutation selected from the group consisting of L235V, F243L, R292P, Y300L, P396L, and a combination thereof, numbered according to the EU numbering system, may be made in the constant region of an antibody described herein. In certain embodiments, a mutation selected from the group consisting of S267E, L328F, and a combination thereof, numbered according to the EU numbering system, may be made in the constant region of an antibody described herein.

[0129] In a specific embodiment, an antibody described herein comprises the constant region of an IgGi with an N297Q or N297A amino acid substitution, numbered according to the EU numbering system. In certain embodiments, an antibody described herein comprises the constant region of an IgGi with a mutation selected from the group consisting of D265A, P329A, and a combination thereof, numbered according to the EU numbering system. In another embodiment,an antibody described herein comprises the constant region of an IgGi with a mutation selected from the group consisting of L234A, L235A, and a combination thereof, numbered according to the EU numbering system. In another embodiment, an antibody described herein comprises the constant region of an IgGi with L234A and L235A mutations, numbered according to the EU numbering system (referred to as “LALA”). In another embodiment, an antibody described herein comprises the constant region of an IgGi with H435R and Y436F mutations, numbered according to the EU numbering system (referred to as “RF”). In another embodiment, an antibody described herein comprises the constant region of an IgGi with a mutation selected from the group consisting of L234A, L235A, H435R, Y436F, and a combination thereof, numbered according to the EU numbering system. In another embodiment, an antibody described herein comprises the constant region of an IgGi with L234A, L235A, H435R, and Y436F mutations, numbered according to the EU numbering system.

[0130] In another embodiment, an antibody described herein comprises the constant region of an IgGi with a mutation selected from the group consisting of L234F, L235F, N297A, and a combination thereof, numbered according to the EU numbering system. In certain embodiments, amino acid residues in the constant region of an antibody described herein in the positions corresponding to positions L234, L235, and D265 in a human IgGi heavy chain, numbered according to the EU numbering system, are not L, L, and D, respectively. This approach is described in detail in International Publication No. WO 14 / 108483, which is herein incorporated by reference in its entirety. In certain embodiments, the amino acids corresponding to positions L234, L235, and D265 in a human IgGi heavy chain are F, E, and A; or A, A, and A, respectively, numbered according to the EU numbering system.

[0131] In certain embodiments, one or more amino acids selected from amino acid residues 356 and 358 in the constant region of an antibody described herein, numbered according to the EU numbering system, can be replaced with a different amino acid residue. In certain embodiments, an antibody described herein comprises the constant region of an IgGi with a mutation selected from the group consisting of D356E, L358M, and a combination thereof, numbered according to the EU numbering system. In another embodiment, an antibody described herein comprises the constant region of an IgGi with D356E and L358M mutations, numbered according to the EU numbering system.

[0132] In certain embodiments, one or more amino acids selected from amino acid residues 329, 331, and 322 in the constant region of an antibody described herein, numbered according to the EU numbering system, can be replaced with a different amino acid residue such that the antibody has altered Clq binding and / or reduced or abolished complement dependent cytotoxicity (CDC). This approach is described in further detail in U. S. Patent No. 6,194,551 (Idusogie et al. which is herein incorporated by reference in its entirety. In certain embodiments, one or more amino acid residues within amino acid positions 231 to 238 in the N-terminal region of the CH2 domain of an antibody described herein are altered to thereby alter the ability of the antibody to fix complement, numbered according to the EU numbering system. This approach is described further in International Publication No. WO 94 / 29351, which is herein incorporated by reference in its entirety. In certain embodiments, the Fc region of an antibody described herein is modified to increase the ability of the antibody to mediate antibody dependent cellular cytotoxicity (ADCC) and / or to increase the affinity of the antibody for an Fey receptor by mutating one or more amino acids (e.g., introducing amino acid substitutions) at the following positions: 238, 239, 248, 249, 252, 254, 255, 256, 258, 265, 267, 268, 269, 270, 272, 276, 278, 280, 283, 285, 286, 289, 290, 292, 293, 294, 295, 296, 298, 301, 303, 305, 307, 309, 312, 315, 320, 322, 324, 326, 327, 328, 329, 330, 331, 333, 334, 335, 337, 338, 340, 360, 373, 376, 378, 382, 388, 389, 398, 414, 416, 419, 430, 434, 435, 437, 438, or 439, numbered according to the EU numbering system. This approach is described further in International Publication No. WO 00 / 42072, which is herein incorporated by reference in its entirety.

[0133] In certain embodiments, an antibody described herein comprises a modified constant region of an IgGi, wherein the modification increases the ability of the antibody to mediate antibody dependent cellular cytotoxicity (ADCC). In certain embodiments, 0.1, 1, or 10 pg / ml of the antibody is capable of inducing cell death of at least 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, or 60% of LY6K-expressing cells within 1, 2, or 3 hours, as assessed by methods described herein and / or known to a person of skill in the art. In certain embodiments, the modified constant region of an IgGi comprises S239D and I332E substitutions, numbered according to the EU numbering system. In certain embodiments, the modified constant region of an IgGi comprises S239D, A330L, and I332E substitutions, numbered according to the EU numbering system. In certain embodiments, the modified constant region of an IgGi comprises L235V, F243L, R292P, Y300L, and P396L substitutions, numbered according to the EU numbering system. In certainembodiments, the antibody is capable of inducing cell death in effector T cells and Tregs, wherein the percentage of Tregs that undergo cell death is higher than the percentage of effector T cells that undergo cell death by at least 1.2 fold, 1.3 fold, 1.4 fold, 1.5 fold, 1.6 fold, 1.7 fold, 1.8 fold, 1.9 fold, 2 fold, 2.5 fold, 3 fold, 3.5 fold, 4 fold, 4.5 fold, or 5 fold.

[0134] In certain embodiments, an antibody described herein comprises the constant region of an IgG4antibody and the serine at amino acid residue 228 of the heavy chain, numbered according to the EU numbering system, is substituted for proline.

[0135] In certain embodiments, any of the constant region mutations or modifications described herein can be introduced into one or both heavy chain constant regions of an antibody described herein having two heavy chain constant regions.Antibody-drug Conjugates

[0136] In another aspect, the instant disclosure provides antibody-drug conjugates (ADCs) of general structure (1)AB-[(L6)b-{Z-L-D}x]y(1)wherein:AB is an antibody that specifically binds to LY6K (e., human LY6K);L is a linker that links Z to D;Z is a connecting group;L6is - GlcNAc(Fuc)w-(G)j-S-(L7)W’-, wherein G is a monosaccharide, j is an integer in the range of 0 - 10, S is a sugar or a sugar derivative, GlcNAc is N-acetylglucosamine and Fuc is fucose, w is 0 or 1, w’ is 0, 1 or 2 and L7is -N(H)C(O)CH2- -N(H)C(O)CF2- or -CH2-;D is selected from the group consisting of taxanes, anthracyclines, camptothecins, epothilones, mytomycins, combretastatins, vinca alkaloids, tubulysins, enediynes (e.g., calicheamicins), amanitins, duocarmycins, maytansinoids, amatoxins, dolastatins, auristatins, eribulins, BCL-XL inhibitors, hemiasterlins, KSP inhibitors, radioisotopes, TLR agonists, indolinobenzodiazepine dimers or pyrrolobenzodiazepine dimers (PBDs), and analogues or prodrugs thereof;b is 0 or 1;x is 1 or 2; andy is 1, 2, 3 or 4.

[0137] Also provided herein are salts, e.g., pharmaceutically acceptable salts, of the ADC according to structure (1).

[0138] In another aspect, provided herein is a process for preparing an ADC provided herein, comprising reacting the compound according to general structure (2) with an antibody (3). The compound according to general structure (2) comprises a reactive moiety Q and the antibody a reactive moiety F which is capable of reacting with Q in a conjugation reaction, wherein Q and F react to form connecting group Z. In this reaction, a conjugate according to general structure (1) is formed. The process according to this aspect this concerns the following bioconjugation reaction:(3) (2) (1)

[0139] The ADC according to structure (1) is defined below. The structural features of the ADC according to structure (1) also apply to the compound according to structure (2) and the antibody according to structure (3), as those are unchanged in the conjugation reaction except for reactive moieties F and Q, which are transformed into connecting group Z upon reaction of the compound according to structure (2) with an antibody according to structure (3).

[0140] As will be understood by the skilled person, the definition of the chemical moieties, as well as their embodiments as described herein, apply to all aspects of the present disclosure. Antibody-drug conjugate of general structure (1)

[0141] In an aspect, provided herein is an ADC of general structure (1):AB-[(L6)b-{Z-L-D}x]y(1)wherein:AB is an antibody that specifically binds to LY6K (e.g., human LY6K);b is 0 or 1;L6is -GlcNAc(Fuc)w-(G)j-S-(L7)W’-, wherein G is a monosaccharide, j is an integer in the range of 0 - 10, S is a sugar or a sugar derivative, GlcNAc is N-acetylglucosamine and Fuc is fucose, w is 0 or 1, w’ is 0, 1 or 2 and L7is -N(H)C(O)CH2-, -N(H)C(O)CF2- or -CH2-;Z is a connecting group;L is a linker that links Z to D;D is selected from the group consisting of taxanes, anthracyclines, camptothecins, epothilones, mytomycins, combretastatins, vinca alkaloids, tubulysins, enediynes (e.g., calicheamicins), amanitins, duocarmycins, maytansinoids, amatoxins, dolastatins, auristatins, eribulins, BCL-XL inhibitors, hemiasterlins, KSP inhibitors, radioisotopes, TLR agonists, indolinobenzodiazepine dimers or pyrrolobenzodiazepine dimers (PBDs), and analogues or prodrugs thereof;x is 1 or 2; andy is 1, 2, 3 or 4.Antibody AB

[0142] The ADC according to the present disclosure comprises an antibody that specifically binds to LY6K (e.g., human LY6K). In certain embodiments, the antibody is any of the anti-LY6K antibodies disclosed herein. In certain embodiments, the antibody comprises: a VH comprising the CDRH1, CDRH2, and CDRH3 amino acid sequences of the VH amino acid sequence set forth in any one of SEQ ID NOs: 1-34; and a VL comprising the CDRL1, CDRL2, and CDRL3 amino acid sequences of the VL amino acid sequence set forth in any one of SEQ ID NOs: 35-68. In certain embodiments, the VH amino acid sequence and VL amino acid sequence are as set forth in: SEQ ID NOs: 1 and 35, 2 and 36, 3 and 37, 4 and 38, 5 and 39, 6 and 40, 7 and 41, 8 and 42, 9 and 43, 10 and 44, 11 and 45, 12 and 46, 13 and 47, 14 and 48, 15 and 49, 16 and 50, 17 and 51, 18 and 52, 19 and 53, 20 and 54, 21 and 55, 22 and 56, 23 and 57, 24 and 58, 25 and 59, 26 and 60, 27 and 61, 28 and 62, 29 and 63, 30 and 64, 31 and 65, 32 and 66, 33 and 67, or 34 and 68, respectively. In certain embodiments, the VH amino acid sequence and VL amino acid sequence are as set forth in: SEQ ID NOs: 2 and 36 or 16 and 50, respectively. In certain embodiments, the antibody comprises the CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in: SEQ ID NOs: 70, 91, 125, 159, WAS, and SEQ ID NO: 199 or SEQ ID NOs: 74, 105, 138, 170, 185, and 211, respectively. In certain embodiments, the antibody comprises the VH amino acid sequence and VL amino acid sequence set forth in SEQ ID NOs: 2 and 36 or 16 and 50, respectively.

[0143] In certain embodiments, the antibody specifically binds to an amino acid sequence selected from the group consisting of SEQ ID NOs: 233-238. In certain embodiments, the antibody specifically binds to the amino acid sequence of SEQ ID NOs: 233 and 239. In certain embodiments, the antibody specifically binds to the amino acid sequence of SEQ ID NOs: 234 and240. In certain embodiments, the antibody specifically binds to the amino acid sequence of SEQ ID NOs: 237 and 247.Linker L6

[0144] In case reactive group F is directly connected to the antibody, or even part of the antibody structure, linker L6that connects AB to F (for antibodies of structure (3)) or AB to Z (for conjugates of structure (1) is absent and b = 0. This is for example the case for cysteine conjugation and lysine conjugation. Alternatively, reactive group F may also be introduced onto the antibody using a linker L6that connects AB to F (for antibodies of structure (3)) or AB to Z (for conjugates of structure (1), in which case L6is present and b = 1. In case L6is present, reactive group F is typically introduced at the glycan of the antibody. This is for example the case for conjugation via an artificially introduced reactive group F, such as for example using transglutaminase or by enzymatic glycan modification (e.g, glycosyltransferase or oc-l,3-mannosyl-glycoprotein-2-P-N-acetylglucosaminyl-transferase). For example, a modified sugar residue S(F)Xmay be introduced at the glycan, extending the glycan with one monosaccharide residue S, which introduces x reactive groups F on the glycan of an antibody. In certain embodiments, conjugation occurs via the glycan of the antibody and b = 1. In certain embodiments, the site of conjugation is at the heavy chain of the antibody.

[0145] If present, L6is a linker that links AB to F or Z, and is represented by -GlcNAc(Fuc)w-(G)j-S-(L7)W- wherein G is a monosaccharide, j is an integer in the range of 0 - 10, S is a sugar or a sugar derivative, GlcNAc is N-acetylglucosamine and Fuc is fucose, w is 0 or 1, w’ is 0, 1 or 2 and L7is -N(H)C(O)CH2-, -N(H)C(O)CF2- or -CH2-. Typically, L6is at least partly formed by the glycan of the antibody. All recombinant antibodies, generated in mammalian host systems, contain the conserved N-glycosylation site at the asparagine residue at or close to position 297 of the heavy chain, which is modified by a glycan of the complex type. In certain embodiments, this naturally occurring glycosylation site of antibodies is used. In certain embodiments, other glycosylation sites, including artificially introduced ones, are used for the connection of linker L6. In certain embodiments, L6is connected to an amino acid of the antibody which is located at a position in the range of 250 - 350 of the heavy chain, e.g., in the range of 280 - 310 of the heavy chain, in the range of 295 - 300 of the heavy chain, or at position 297 of the heavy chain.

[0146] The -GlcNAc(Fuc)w-(G)j- of L6is the glycan of the antibody, or part thereof. The -GlcNAc(Fuc)b-(G)j- of the glycan thus typically originates from the original antibody, whereinGlcNAc is an N-acetylglucosamine moiety and Fuc is a fucose moiety. Fuc is typically bound to GlcNAc via an oc-l,6-glycosidic bond. Normally, antibodies may (w = 1) or may not be fucosylated (w = 0). In the context of the present invention, the presence of a fucosyl moiety is irrelevant, and similar effects are obtained with fucosylated (w = 1) and non-fucosylated (w = 0) antibody conjugates. The GlcNAc residue may also be referred to as the core-GlcNAc residue and is the monosaccharide that is directly attached to the peptide part of the antibody.

[0147] S may be directly connected to the core-GlcNAc(Fuc)wmoiety, i.e., j = 0, meaning that the remainder of the glycan is removed from the core-GlcNAc(Fuc)wmoiety before S is attached. Such trimming of glycans is well-known in the art and can be achieved by the action of an endoglycosidase. Alternatively, there are one or more monosaccharide residues present in between the core-GlcNAc(Fuc)wmoiety and S, i.e., j is an integer in the range of 1 - 10, e.g., j = 2 - 5. In certain embodiments, (G)j is an oligosaccharide fraction comprising j monosaccharide residues G, wherein j is an integer in the range of 2 - 5. (G)j is connected to the GlcNAc moiety of GlcNAc(Fuc)w, typically via a P-1,4 bond. In certain embodiments, j is 3, 4 or 5. Although any monosaccharide that may be present in a glycan may be employed as G, in certain embodiments Gis individually selected from the group consisting of galactose, glucose, N-acetylgalactosamine, N-acetylglucosamine, mannose and N-acetylneuraminic acid. In certain embodiments, G is individually selected from galactose, N-acetylglucosamine, and mannose. In certain embodiments, ADCs having j below 4 show no or hardly any binding to the Fc-gamma receptor, while antibody conjugates having j in the range of 4 - 10 do bind to the Fc-gamma receptor. Thus, in certain embodiments, by selecting a certain value for j, the desired extent of binding to the Fc-gamma receptor can be obtained. In certain embodiments, j = 0, 3, 4, 5, 6, 7, 8, 9 or 10. In certain embodiments, j = 0, 3, 4 or 5. In certain embodiments, the antibody is trimmed and j = 0.

[0148] S is a sugar or sugar derivative. The term “sugar derivative” is herein used to indicate a derivative of a monosaccharide sugar, i.e., a monosaccharide sugar comprising substituents and / or functional groups. Suitable examples for S include glucose (Glc), galactose (Gal), mannose (Man), fucose (Fuc), amino sugars and sugar acids, e.g., glucosamine (GlcNFF), galactosamine (GalNFF) N-acetylglucosamine (GlcNAc), N-acetylgalactosamine (GalNAc), sialic acid (Sia) which is also referred to as N-acetylneuraminic acid (NeuNAc), and N-acetylmuramic acid (MurNAc), glucuronic acid (GlcA) and iduronic acid (IdoA). In certain embodiments, S is selected from Glc, Gal, GlcNAc and GalNAc. In certain embodiments, S is GalNAc.

[0149] x is an integer that denotes the number of connecting groups Z (for conjugate (1)) or reactive groups F (for antibody (3)) that are attached to sugar (derivative) S. Thus, the antibody of the ADC provided herein comprises a moiety S comprising x reactive moieties F. Each of these reactive moieties F are reacted with a reactive moiety Q of the compound according to general structure (2), such that x connecting groups Z are formed and x compounds according to general structure (2) are attached to a single occurrence of S. x is 1 or 2, e.g., x = 1.

[0150] Connecting group Z (for conjugate (1)) or reactive group F (for antibody (3)) may be attached directly to S, or there may be a linker L7present in between S and Z or F. L7is a linker that links S with Z. L7may be present (w’ = 1 or 2) or absent (w’ = 0). Typically, each moiety Z may be connected to S via a linker L7, thus in one embodiment w’ = 0 of x. In certain embodiments, L7is absent and each connecting moiety Z is directly attached to S. If present, L7may be selected from -N(H)C(O)CH2-, -N(H)C(O)CF2- or -CH2-. In certain embodiments, x = 1 and w’ = 0 or 1, e.g., x = 1 and w’ = 0.

[0151] y is an integer that denotes the number of sugar(s) (derivative(s)) S, each having x reactive groups F or connected to x connecting groups Z, that are connected to the antibody. In certain embodiments, y is 1, 2, 3 or 4. In certain embodiments, y = 2 or 4. In certain embodiments, y = 2. Thus, the antibody comprises y moieties S, each of which comprises x reactive moieties F. Each of these reactive moieties F are reacted with reactive moiety Q of the compound according to general structure (2), such that x + y connecting groups Z are formed and x + y compounds according to general structure (2) are attached to a single antibody. Each compound according to general structure (2) may comprise multiple payloads, e.g., by virtue of branching nitrogen atom N* in L. In certain embodiments, each compound according to general structure (2) comprises 1 or 2 occurrences of D. In certain embodiments, each compound according to general structure (2) comprises 2 occurrences ofD. In certain embodiments, linker L1comprises a branching nitrogen atom N* to which a second occurrence of D is connected.

[0152] The amount of payload (D) molecules attached to a single antibody is known in the art as the DAR (drug-antibody ratio). In certain embodiments of the ADCs provided herein, the DAR is an integer in the range 1 - 8. In certain embodiments, the DAR is 2 or 4. In certain embodiments, the DAR is 4. In certain embodiments, the DAR is an integer in the range (x + y) to [(x + y)x2], In certain embodiments, the DAR = [(x + y) x 2], In certain embodiments, the values for x and y are 1 and 2, respectively, and the DAR is 4. It will be appreciated that these are theoretical DARvalues, and in practice the DAR may slightly deviate from this value, by virtue of incomplete conjugation. Typically, the conjugates are obtained as a stochastic mixture of antibody-drug conjugates, with DAR values varying between individual conjugates, and depending on the conjugation technique used the DAR may have a broad distribution (e.g., DAR = 0 - 10) or a narrow distribution (e.g., DAR = 3 - 4). In case of such mixture, DAR often refers to the average DAR of the mixture. This is well-known in the art of bioconjugation. In certain embodiments of the ADCs provided herein, the conjugation occurs via the glycan (i.e., b = 1 and L6is present), and the ADCs have a close-to-theoretical DAR. For example, when the theoretical DAR is 4, DAR values above 3.6 or even above 3.8 are readily obtained, indicating that most antibodies in the reaction mixture have reacted completely and have a DAR of 4.Connecting group Z

[0153] Z is a connecting group, which covalently connects both parts of the ADC according to the present disclosure. The term “connecting group” herein refers to the structural element, resulting from the reaction between Q and F, connecting one part of the conjugate with another part of the same conjugate. As will be understood by the person skilled in the art, the nature of a connecting group depends on the type of reaction with which the connection between the parts of said compound is obtained. As an example, when the carboxyl group of R-C(O)-OH is reacted with the amino group of H2N-R’ to form R-C(0)-N(H)-R’, R is connected to R’ via connecting group Z, and Z may be represented by the group -C(O)-N(H)-. Since connecting group Z originates from the reaction between Q and F, it can take any form.

[0154] Since more than one reactive moiety F can be present or introduced in an antibody, an ADC provided herein may comprise per biomolecule more than one payload D, e.g., 1 - 8 payloads D; 1, 2, 3 or 4 payloads D; or 2 or 4 payloads D. The number of payloads is typically an even integer, in view of the symmetric nature of antibodies. In other words, when one side of the antibody is functionalized with F, the symmetrical counterpart will also be functionalized. Alternatively, in case naturally occurring thiol groups of the cysteine residues of a protein are used as F, the value of m can be anything and may vary between individual conjugates.

[0155] In a compound according to structure (1), connecting group Z connects D via linker L to AB, optionally via L6. Numerous reactions are known in the art for the attachment of a reactive group Q to a reactive group F. Consequently, a wide variety of connecting groups Z may be present in the conjugate according to the invention. In one embodiment, the reactive group Q is selectedfrom the options described above. In certain embodiments, the reactive group Q is selected from the options depicted in FIG. 1, and complementary reactive groups F and the thus obtained connecting groups Z are known to a person skilled in the art. Several examples of suitable combinations of F and Q, and of connecting group Z3that will be present in a bioconjugate when a linker-conjugate comprising Q is conjugated to a biomolecule comprising a complementary reactive group F, are shown in FIG. 1.

[0156] In certain embodiments, when F comprises or is a thiol group, complementary groups Q include N-maleimidyl groups and alkenyl groups, and the corresponding connecting groups Z are as shown in FIG. 1. When F comprises or is a thiol group, complementary groups Q also include allenamide groups.

[0157] In certain embodiments, when F comprises or is an amino group, complementary groups Q include ketone groups and activated ester groups, and the corresponding connecting groups Z are as shown in FIG. 1.

[0158] In certain embodiments, when F comprises or is a ketone group, complementary groups Q include (O-alkyl)hydroxylamino groups and hydrazine groups, and the corresponding connecting groups Z are as shown in FIG. 1.

[0159] In certain embodiments, when F comprises or is an alkynyl group, complementary groups Q include azido groups, and the corresponding connecting group Z is as shown in FIG. 1.

[0160] In certain embodiments, when F comprises or is an azido group, complementary groups Q include alkynyl groups, and the corresponding connecting group Z is as shown in FIG. 1.

[0161] In certain embodiments, when F comprises or is a cyclopropenyl group, a transcyclooctene group or a cyclooctyne group, complementary groups Q include tetrazinyl groups, and the corresponding connecting group Z is as shown in FIG. 1. In these particular cases, Z is only an intermediate structure and will expel N2, thereby generating a dihydropyridazine (from the reaction with alkene) or pyridazine (from the reaction with alkyne).

[0162] Additional suitable combinations of F and Q, and the nature of resulting connecting group Z3 are known to a person skilled in the art, and are described, e.g., in G. T. Hermanson, “Bioconjugate Techniques”, Elsevier, 3rd Ed. 2013 (ISBN:978-0-12-382239-0), in particular in Chapter 3, pages 229 - 258, incorporated herein by reference. A list of complementary reactive groups suitable for bioconjugation processes is disclosed in Table 3.1, pages 230 - 232 of Chapter3 of G. T. Hermanson, “Bioconjugate Techniques”, Elsevier, 3rd Ed. 2013 (ISBN:978-0-12-382239-0), and the content of this Table is expressly incorporated by reference herein.

[0163] In certain embodiments, connecting group Z is obtained by a cycloaddition or a nucleophilic reaction. In certain embodiments, the cycloaddition is a [4+2] cycloaddition or a 1,3-dipolar cycloaddition or the nucleophilic reaction is a Michael addition or a nucleophilic substitution. Such a cycloaddition or nucleophilic reaction occurs via a reactive group F, connected to S, and reactive group Q, connected to D via L. Conjugation reactions via cycloadditions or nucleophilic reactions are known to the skilled person, and the skilled person is capable of selecting appropriate reaction partners F and Q, and will understand the nature of the resulting connecting group Z.

[0164] In certain embodiments, Z is formed by a cycloaddition. In certain embodiments, the cycloaddition is a (4+2)-cycloaddition (e.g, a Diels-Alder reaction) or a (3+2)-cycloaddition (e.g, a 1,3-dipolar cycloaddition). In certain embodiments, the conjugation is the Diels-Alder reaction or the 1,3-dipolar cycloaddition. In certain embodiments, the Diels-Alder reaction is the inverseelectron demand Diels- Alder cycloaddition. In certain embodiments, the 1,3-dipolar cycloaddition is used. In certain embodiments, the alkyne-azide cycloaddition is used. In certain embodiments, Q is or comprises an alkyne group and F is an azido group. Cycloadditions, such as Diels-Alder reactions and 1,3-dipolar cycloadditions are known in the art, and the skilled person knowns how to perform them.

[0165] In certain embodiments, Z comprises a moiety selected from the group consisting of a triazole, a cyclohexene, a cyclohexadiene, a [2.2.2]-bicyclooctadiene, a [2.2.2]-bicyclooctene, an isoxazoline, an isoxazolidine, a pyrazoline, a piperazine, a thioether, an amide or an imide group. In certain embodiments, Z comprises a triazole moiety. In certain embodiments, Z comprises a (hetero)cycloalkene moiety, i.e., formed from Q comprising a (hetero)cycloalkyne moiety. In certain embodiments, Z comprises a (hetero)cycloalkane moiety, i.e., formed from Q comprising a (hetero)cycloalkene moiety. In certain embodiments, Z has the structure (Z 1):wherein the bond depicted as - is a single bond or a double bond. Furthermore: ring Z is obtained by a cycloaddition, e.g., ring Z is selected from (Za) - (Zj) defined below, wherein the carbon atoms labelled with ** correspond to the two carbon atoms of the bond depicted as - of (Zl) to which ring Z is fused;R15is independently selected from the group consisting of hydrogen, halogen, -OR16, -NO2, -CN, -S(O)2R16, -S(O)3(’), CI - C24 alkyl groups, Ce - C24 (hetero)aryl groups, C7 - C24 alkyl(hetero)aryl groups and C7 - C24 (het ero)aryl alkyl groups and wherein the alkyl groups, (hetero)aryl groups, alkyl(hetero)aryl groups and (hetero)arylalkyl groups are optionally substituted, wherein two substituents R15may be linked together to form an optionally substituted annulated cycloalkyl or an optionally substituted annulated (hetero)arene substituent, and wherein R16is independently selected from the group consisting of hydrogen, halogen, Ci - C24 alkyl groups, Ce - C24 (hetero)aryl groups, C7 - C24 alkyl(hetero)aryl groups and C7 - C24 (hetero)aryl alkyl groups;Y2is C(R31)2, O, S, S( I )R31, S(O)R31, S(O)=NR31or NR31, wherein S( l )is a cationic sulphur atom counterbalanced by B, wherein B( )is an anion, and wherein each R31individually is R15or a connection with D, connected via L;u is 0, 1, 2, 3, 4 or 5;u’ is 0, 1, 2, 3, 4 or 5, wherein u + u’ = 0, 1, 2, 3, 4, 5, 6, 7 or 8;v = an integer in the range 8 - 16; andRing A is formed by the cycloaddition, e.g., Ring A is selected from (Za) - (Zj).

[0166] In certain embodiments, the bond depicted as - is a double bond, and u + u’ — 4, 5, 6, 7 or 8. In certain embodiments, the wavy bond labelled with * is connected to S and the wavy bond labelled with ** is connected to L.

[0167] In certain embodiments, Z comprises a (hetero)cycloalkene moiety, i.e., the bond depicted as - is a double bond. In certain embodiments, Z is selected from the structures (Z2) - (Z20), depicted here below:(Z16) (Z17) (Z18) (Z19) (Z20) wherein the connection to L is depicted with the wavy bond. B(-1is an anion, e.g., a pharmaceutically acceptable anion. Ring Z is formed by the cycloaddition reaction, and is, in certain embodiments, a triazole, a cyclohexene, a cyclohexadiene, a [2.2.2]-bicyclooctadiene, a [2.2.2]-bicyclooctene, an isoxazoline, an isoxazolidine, a pyrazoline or a piperazine. In certain embodiments, ring Z is a triazole ring. Ring Z may have the structure selected from (Za) - (Zm) depicted below, wherein the carbon atoms labelled with ** correspond to the two carbon atoms of the (hetero)cycloalkane ring of (Z2) - (Z20) to which ring Z is fused. In certain embodiments, ring Z is selected from (Za) - (Zj). In certain embdiments, ring Z is selected from (Za), (Zd) and (Zh). In certain embodiments, ring Z has structure (Za). Since the connecting group Z is formed by reaction with a (hetero)cycloalkyne in the context of the present embodiment, the bound depicted above as - is a double bond.

[0168] In certain embodiments, Z is selected from the structures (Z21) - (Z38) and (Z38a), depicted here below:(Z21) (Z22) (Z23) (Z24) (Z25) (Z26)(Z35) (Z36) (Z37) (Z38) (Z38a)wherein the connection to L is depicted with the wavy bond. In structureis an anion, e.g., a pharmaceutically acceptable anion. In certain embodiments, Ring Z is selected from structures (Za) - (Zm), as defined above. In certain embodiments, Ring Z is selected from structures (Za) - (Zj), as defined above.

[0169] In certain embodiments, Z comprises a (hetero)cyclooctene moiety or a (hetero)cycloheptene moiety, e.g., according to structure (Z8), (Z26), (Z27), (Z28) or (Z37) or (Z38a), or according to structure (Z8), (Z26), (Z27), (Z28) or (Z37), which are optionally substituted. Each of these options for Z are further defined here below.

[0170] Thus, in certain embodiments, Z comprises a heterocycloheptene moiety according to structure (Z37), which is optionally substituted. In certain embodiments, the heterocycloheptyne moiety according to structure (Z37) is not substituted.

[0171] In certain embodiments, Z comprises a (hetero)cyclooctene moiety according to structure (Z8), which is optionally substituted. In certain embodiments, Z comprises a (hetero)cyclooctene moiety according to structure (Z29), which is optionally substituted. In certain embodiments, the cyclooctene moiety according to structure (Z8) or (Z29) is not substituted. Incertain embodiments, Z comprises a (hetero)cyclooctene moiety according to structure (Z39) as shown below, wherein V is (CH2)I and 1 is an integer in the range of 0 to 10, e.g., in the range of 0 to 6. In certain embodiments, 1 is 0, 1, 2, 3 or 4. In certain embodiments, 1 is 0, 1 or 2. In certain embodiments, 1 is 0 or 1. In certain embodiments, in the context of group (Z39), 1 is 1. In certain embodiments, Z is according to structure (Z42), defined further below.

[0172] In certain embodiments, Z comprises a (hetero)cyclooctene moiety according to structure (Z26), (Z27) or (Z28), which are optionally substituted. In certain embodiments, Z comprises a (hetero)cyclooctene moiety according to structure (Z40) or (Z41) as shown below, wherein Y1is O or NR11, wherein R11is independently selected from the group consisting of hydrogen, a linear or branched Ci - C12 alkyl group or a C4 - C12 (hetero)aryl group. The aromatic rings in (Z40) are optionally O-sulfonylated at one or more positions, whereas the rings of (Z41) may be halogenated at one or more positions. In certain embodiments, the (hetero)cyclooctene moiety according to structure (Z40) or (Z41) is not further substituted. In certain embodiments, Z is according to structure (Z43), defined further below.

[0173] In certain embodiments, Z comprises a heterocycloheptenyl group and is according to structure (Z37).(Z37) (Z39) (Z40) (Z41)

[0174] In certain embodiments, Z comprises a cyclooctenyl group and is according to structure (Z42):wherein:the bond labelled with * is connected to S and the wavy bond labelled with ** is connected to L;R13is independently selected from the group consisting of hydrogen, halogen, -OR16, -NO2, -CN, -S(O)2R16, -S(O)3(_:), CI - C24 alkyl groups, C5 - C24 (hetero)aryl groups, C7 - C24 alkyl(hetero)aryl groups and C7 - C24 (hetero)aryl alkyl groups and wherein the alkyl groups, (hetero)aryl groups, alkyl(hetero)aryl groups and (hetero)arylalkyl groups are optionally substituted, wherein two substituents R15may be linked together to form an optionally substituted annulated cycloalkyl or an optionally substituted annulated (hetero)arene substituent, and wherein R16is independently selected from the group consisting of hydrogen, halogen, Ci - C24 alkyl groups, Ce - C24 (hetero)aryl groups, C7 - C24 alkyl(hetero)aryl groups and C7 - C24 (hetero)aryl alkyl groups;R18is independently selected from the group consisting of hydrogen, halogen, Ci - C24 alkyl groups, Ce - C24 (hetero)aryl groups, C7 - C24 alkyl(hetero)aryl groups and C7 - C24 (hetero)arylalkyl groups;R19is selected from the group consisting of hydrogen, halogen, Ci - C24 alkyl groups, Ce - C24 (hetero)aryl groups, C7 - C24 alkyl(hetero)aryl groups and C7 - C24 (hetero)arylalkyl groups, the alkyl groups optionally being interrupted by one of more hetero-atoms selected from the group consisting of O, N and S, wherein the alkyl groups, (hetero)aryl groups, alkyl(hetero)aryl groups and (hetero)arylalkyl groups are independently optionally substituted, or R19is a second occurrence of Z (or Q) or D connected via a spacer moiety; and1 is an integer in the range 0 to 10.

[0175] In certain embodiments of the group according to structure (Z42), R13is independently selected from the group consisting of hydrogen, halogen, -OR16, Ci - Ce alkyl groups, C5 - Ce (hetero)aryl groups, wherein R16is hydrogen or Ci - Ce alkyl. In certain embodiments, R15is independently selected from the group consisting of hydrogen and Ci - Ce alkyl. In certain embodiments, all R15are H. In certain embodiments of the group according to structure (Z42), R18is independently selected from the group consisting of hydrogen, Ci - Ce alkyl groups. In certain embodiments, both R18are H. In certain embodiments of the group according to structure (Z42), R19is H. In certain embodiments of the group according to structure (Z42), I is 0 or 1. In certain embodiments, 1 is 1.

[0176] In certain embodiments, Z comprises a (hetero)cyclooctynyl group and is according to structure (Z43):wherein:the bond labelled with * is connected to S and the wavy bond labelled with ** is connected to L;R15is independently selected from the group consisting of hydrogen, halogen, -OR16, -NO2, -CN, -S(O)2R16, -S(O)3(-), CI - C24 alkyl groups, C5 - C24 (hetero)aryl groups, C7 - C24 alkyl(hetero)aryl groups and C7 - C24 (hetero)aryl alkyl groups and wherein the alkyl groups, (hetero)aryl groups, alkyl(hetero)aryl groups and (hetero)arylalkyl groups are optionally substituted, wherein two substituents R13may be linked together to form an optionally substituted annulated cycloalkyl or an optionally substituted annulated (hetero)arene substituent, and wherein R16is independently selected from the group consisting of hydrogen, halogen, Ci - C24 alkyl groups, Ce - C24 (hetero)aryl groups, C7 - C24 alkyl(hetero)aryl groups and C7 - C24 (hetero)aryl alkyl groups;Y is N or CR15.

[0177] In certain embodiments of the group according to structure (Z43), R15is independently selected from the group consisting of hydrogen, halogen, -OR16, -S(O)3( ), Ci - Ce alkyl groups, C5 - Ce (hetero)aryl groups, wherein R16is hydrogen or Ci - Ce alkyl. In certain embodiments, R15is independently selected from the group consisting of hydrogen and -S(O)3(-). In certain embodiments of the group according to structure (Z43), Y is N or CH. In certain embodiments, Y = N.

[0178] In certain embodiments, Z comprises a heterocycloheptynyl group and is according to structure (Z37) or (Z38a), wherein ring Z is a triazole. In certain embodiments, Z comprises a heterocycloheptynyl group and is according to structure (Z37), wherein ring Z is a triazole.(Z37) (Z38a)

[0179] In certain embodiments, Z comprises a (hetero)cycloalkane moiety, i.e., the bond depicted as - is a single bond. The (hetero)cycloalkane group may also be referred to as a heterocycloalkanyl group or a cycloalkanyl group, e.g., a cycloalkanyl group, wherein the (hetero)cycloalkanyl group is optionally substituted. In certain embodiments, the (hetero)cycloalkanyl group is a (hetero)cyclopropanyl group, a (hetero)cyclobutanyl group, a norbornane group, a norbornene group, a (hetero)cycloheptanyl group, a (hetero)cyclooctanyl group, a (hetero)cyclononnyl group or a (hetero)cyclodecanyl group, which may all optionally be substituted. In certain embodiments, the (hetero)cycloalkanyl group is a (hetero)cyclopropanyl group, a (hetero)cycloheptanyl group, or a (hetero)cyclooctanyl group, wherein the (hetero)cyclopropanyl group, the trans-(hetero)cycloheptanyl group or the (hetero)cyclooctanyl group is optionally substituted. In certain embodiments, Z comprises a cyclopropanyl moiety according to structure (Z44), a hetereocyclobutane moiety according to structure (Z45), a norbornane or norbornene group according to structure (Z46), a (hetero)cycloheptanyl moiety according to structure (Z47) or a (hetero)cyclooctanyl moiety according to structure (Z48). In certain embodiments, Y3is selected from C(R23)2, NR23or O, wherein each R23is individually hydrogen, Ci - Ce alkyl or is connected to L, optionally via a spacer, and the bond labelled -is a single or double bond. In certain embodiments, the cyclopropanyl group is according to structure (Z49). In certain embodiments, the (hetero)cycloheptane group is according to structure (Z50) or (Z51). In certain embodiments, the (hetero)cyclooctane group is according to structure (Z52), (Z53), (Z54), (Z55) or (Z56).(Z53) (Z54) (Z55) (Z56)wherein the R group(s) on Si in (Z50) and (Z51) are typically alkyl or aryl, e.g., Ci-Ce alkyl. Ring Z is typically selected from structures (Zn) - (Zu), wherein the carbon atoms labelled with ** correspond to the two carbon atoms of the (hetero)cycloalkane ring of (Z44) - (Z56) to which ring Z is fused, and the carbon a carbon labelled with * is directly connected to the peptide chain of the antibody. In certain embodiments, ring Z is selected from (Zo) - (Zr). Since the connecting group Z is formed by reaction with a (hetero)cycloalkene in the context of the present embodiment, the bound depicted above as - is a single bond.

[0180] In certain embodiments, Z is formed by a nucleophilic reaction. In certain embodiments, Z is formed by a nucleophilic substitution. In certain embodiments, Z is formed by a Michael addition. In certain embodiments, the Michael reaction is the thiol-maleimide ligation.In certain embodiments, Q is maleimide and F is a thiol group. In certain embodiments, the thiol is present in the sidechain of a cysteine residue. In certain embodiments, connection group Z comprises a succinimidyl ring or its ring-opened succinic acid amide derivative. In certain embodiments, connection group Z comprises a moiety selected from (Z57) - (Z71) depicted here below.wherein, the wavy bond(s) labelled with an * is connected to the antibody Ab, optionally via a linker, and the wavy bond without label to the payload, optionally via a linker. In certain embodiments, R29is C1-12 alkyl. In certain embodiments, R29is Ci-4 alkyl. In certain embodiments, R29is ethyl. In certain embodiments, X1is O or S. In certain embodiments, X1= O. The nitrogen atom labelled with ** in (Z67)-(Z71) corresponds to the nitrogen atom of the side chain of a lysine residue of the antibody. The carbon atoms of the phenyl group of (Z69) and (Z70) are optionally substituted, e.g., optionally fluorinated.

[0181] In certain embodiments, connection group Z comprise a moiety selected from (Zl) - (Z71).Linker L

[0182] Linker L connects pay load D with connecting group Z (in the ADC according to structure (1)) or connects payload D with reactive group Q (in the compound according to structure (2)). Linkers are known in the art and may be cleavable or non-cleavable. In certain embodiments, Linker L comprises a self-immolative group or cleavable linker, comprising a peptide spacer and a para-aminobenzyloxycarbonyl (PABC) moiety or derivative thereof.

[0183] In certain embodiments, linker L is the structure -(L1)n-(L2)o-(L3)p-(L4)q- wherein (L4)qis connect to payload D and (L')nis connected to Z or Q; L1, L2, L3and L4are linkers or linking units and each of n, o, p and q are individually 0 or 1, and wherein n + o + p + q is at least 1. In certain embodiments, at least linkers L1and L2are present (i.e., n= l; o = l; p = 0 or l; q = 0 or 1). In certain embodiments, linkers L1, L2and L3are present and L4is either present or not (i.e., n = 1; o = 1; p = 1; q = 0 or 1). In certain embodiments, linkers L1, L2, L3and L4are present (i.e., n = 1; o = 1; p = 1; q = 1). In certain embodiments, linkers L1, L2and L3are present and L4is not (i.e., n = 1; o = 1; p = 1; q = 0).

[0184] A linker, especially linker L1, may comprise one or more branch-points for attachment of multiple payloads to a single connecting group. In certain embodiments of the ADCs provided herein, the linker of the ADC comprises a branching moiety. A “branching moiety” in the context of the present disclosure refers to a moiety that is embedded in a linker connecting three moieties. In other words, the branching moiety comprises at least three bonds to other moieties, typically one bond connecting to Z or Q, one bond to the payload D and one bond to a second payload D. In certain embodiments, the branching moiety, if present, is embedded in linker L1. In certain embodiments, the branching moiety, if present, is part of Sp2or is the nitrogen atom of NR13. Any moiety that comprises at least three bonds to other moieties is suitable as branching moiety in the context of the present disclosure. In certain embodiments, the branching moiety is selected from a carbon atom, a nitrogen atom, a phosphorus atom, a (hetero)aromatic ring, a (hetero)cycle or a polycyclic moiety. In certain embodiments, the branching moiety is a nitrogen atom.Linker LI

[0185] Linker L1is either absent (n = 0) or present (n = 1). In certain embodiments, linker L1is present and n = 1. In certain embodiments, L1is selected from the group consisting of linear or branched C1-C200 alkylene groups, C2-C200 alkenylene groups, C2-C200 alkynylene groups, C3-C200 cycloalkylene groups, C5-C200 cycloalkenylene groups, C8-C200 cycloalkynylene groups, C7-C200 alkylarylene groups, C7-C200 arylalkylene groups, C8-C200 arylalkenylene groups, C9-C200 arylalkynylene groups. Optionally the alkylene groups, alkenylene groups, alkynylene groups, cycloalkylene groups, cycloalkenyl ene groups, cycloalkynylene groups, alkylarylene groups, arylalkylene groups, arylalkenylene groups and arylalkynylene groups may be substituted, and optionally said groups may be interrupted by one or more heteroatoms, e.g., 1 to 100 heteroatoms. In certain embodiments, said heteroatoms are selected from the group consisting of O, S(O)y- and NR21, wherein y’ is 0, 1 or 2, e.g., y’ = 2, and R21is independently selected from the group consisting of hydrogen, halogen, Ci - C24 alkyl groups, Ce - C24 (hetero)aryl groups, C7 - C24 alkyl(hetero)aryl groups and C7 - C24 (hetero)arylalkyl groups.

[0186] In certain embodiments, linker L1comprises a polar group. Such a polar group may be selected from (poly)ethylene glycoldiamines e.g., l,8-diamino-3,6-dioxaoctane or equivalents comprising longer ethylene glycol chains), (poly)ethylene glycol or (poly)ethylene oxide chains, (poly)propylene glycol or (poly)propylene oxide chains and l,z’ -diaminoalkanes (wherein z’ is the number of carbon atoms in the alkane, e.g., z’ = 1 - 10), -(O)a-C(O)-NH-S(O)2-NR13- (as further defined below, see structure (23)), -C(S(O)3(')-, -C(C(O)2(-))-, -S(O)2-, -P(O)2f-O(CH2CH2O)t-, -NR30(CH2CH2NR30)t-, and the following two structures:

[0187] The polar group may also comprise an amino acid, e.g., selected from Arg, Glu, Asp, Ser and Thr, wherein a and R13are further defined below for structure (23). In certain embodiments t is an integer in the range of integer in the range of 0 - 15. In certain embodiments, t is an integer in the range of 1 - 10. In certain embodiments, t is an integer in the range of 2 - 5. In certain embodiments, t = 2 or 4. Each R30is individually H, C1.12 alkyl, C1-12 aryl, C1-12 alkaryl or C1-12 aralkyl. Linker L1may comprise more than one such polar group, such as at least two polar groups.The polar group may also be present in a branch of linker L1, which branches off a branching moiety as defined elsewhere. In certain embodiments, a nitrogen or carbon atom is used as branching moiety. In certain embodiments, a -O(CH2CH2O)t- polar group is present in a branch.

[0188] In certain embodiments, Linker L1is or comprises a sulfamide group, e.g., a sulfamide group according to structure (23):

[0189] The wavy lines represent the connection to the remainder of the compound, typically to Q and L2, L3, L4or D, e.g., to Q and L2. In certain embodiments, the (O)aC(O) moiety is connected to Q and the NR13moiety to L2, L3, L4or D, e.g., to L2.

[0190] In structure (23), a = 0 or 1, e.g., a = 1, and R13is selected from the group consisting of hydrogen, Ci - C24 alkyl groups, C3 - C24 cycloalkyl groups, C2 - C24 (hetero)aryl groups, C3 -C24 alkyl(hetero)aryl groups and C3 - C24 (hetero)arylalkyl groups, the Ci - C24 alkyl groups, C3 - C24 cycloalkyl groups, C2 - C24 (hetero)aryl groups, C3 - C24 alkyl(hetero)aryl groups and C3 -C24 (hetero)arylalkyl groups optionally substituted and optionally interrupted by one or more heteroatoms selected from O, S and NR14wherein R14is independently selected from the group consisting of hydrogen and Ci - C4 alkyl groups, or R13is D connected to N via a spacer moiety, e.g., Sp2as defined below. In certain embodiments, D is connected to N via -(B)e-(A)f-(B)g-C(O)-.

[0191] In certain embodiments, R13is hydrogen or a Ci - C20 alkyl group. In certain embodiments, R13is hydrogen or a Ci - Ci6 alkyl group. In certain embodiments, R13is hydrogen or a Ci - C10 alkyl group. In certain embodiments, the alkyl group is optionally substituted and optionally interrupted by one or more heteroatoms selected from O, S and NR14, e.g., O, wherein R14is independently selected from the group consisting of hydrogen and Ci - C4 alkyl groups. In certain embodiments, R13is hydrogen. In certain embodiments, R13is a Ci - C20 alkyl group. In certain embodiments, R13is a Ci - Ci6 alkyl group. In certain embodiments, R13is a Ci - C10 alkyl group. In certain embodiments, the alkyl group is optionally interrupted by one or more O-atoms, and / or the alkyl group is optionally substituted with an -OH group, e.g., a terminal -OH group. In certain embodiments, R13is a (poly)ethylene glycol chain comprising a terminal -OH group. In certain embodiments, R13is selected from the group consisting of hydrogen, methyl, ethyl, n-propyl, i-propyl, n-butyl, s-butyl and t-butyl. In certain embodiments, R13is selected from the group consisting of hydrogen, methyl, ethyl, n-propyl and i-propyl. In certain embodiments, R13is selected from the group consisting of hydrogen, methyl and ethyl. In certain embodiments, R13is hydrogen or methyl. In certain embodiments, R13is hydrogen.

[0192] In certain embodiments, L1is according to structure (24):wherein a and R13are as defined above, Sp1and Sp2are independently spacer moieties and b and c are independently 0 or 1. In certain embodiments, b = 0 or 1 and c = 1. In certain embodiments, b = 0 and c = 1. In certain embodiments, spacers Sp1and Sp2are independently selected from the group consisting of linear or branched C1-C200 alkylene groups, C2-C200 alkenylene groups, C2-C200 alkynylene groups, C3-C200 cycloalkylene groups, C5-C200 cycloalkenylene groups, C8-C200 cycloalkynylene groups, C7-C200 alkylarylene groups, C7-C200 arylalkylene groups, C8-C200 arylalkenylene groups and C9-C200 arylalkynylene groups, the alkylene groups, alkenylene groups, alkynylene groups, cycloalkylene groups, cycloalkenylene groups, cycloalkynylene groups, alkylarylene groups, arylalkylene groups, arylalkenylene groups and arylalkynylene groups being optionally substituted and optionally interrupted by one or more heteroatoms selected from the group of O, S and NR16, wherein R16is independently selected from the group consisting of hydrogen, Ci - C24 alkyl groups, C2 - C24 alkenyl groups, C2 - C24 alkynyl groups and C3 - C24 cycloalkyl groups, the alkyl groups, alkenyl groups, alkynyl groups and cycloalkyl groups being optionally substituted. In certain embodiments, when the alkylene groups, alkenylene groups, alkynylene groups, cycloalkylene groups, cycloalkenylene groups, cycloalkynylene groups, alkylarylene groups, arylalkylene groups, arylalkenylene groups and arylalkynylene groups are interrupted by one or more heteroatoms as defined above, said groups are interrupted by one or more O-atoms, and / or by one or more S-S groups.

[0193] In certain embodiments, spacer moieties Sp1and Sp2, if present, are independently selected from the group consisting of linear or branched C1-C100 alkylene groups, C2-C100 alkenylene groups, C2-C100 alkynylene groups, C3-C100 cycloalkylene groups, C5-C100 cycloalkenylene groups, C8-C100 cycloalkynylene groups, C7-C100 alkylarylene groups, C7-C100 arylalkylene groups, C8-C100 arylalkenylene groups and C9-C100 arylalkynylene groups, the'llalkylene groups, alkenylene groups, alkynylene groups, cycloalkylene groups, cycloalkenylene groups, cycloalkynylene groups, alkylarylene groups, arylalkylene groups, arylalkenylene groups and arylalkynylene groups being optionally substituted and optionally interrupted by one or more heteroatoms selected from the group of O, S and NR16, wherein R16is independently selected from the group consisting of hydrogen, Ci - C24 alkyl groups, C2 - C24 alkenyl groups, C2 - C24 alkynyl groups and C3 - C24 cycloalkyl groups, the alkyl groups, alkenyl groups, alkynyl groups and cycloalkyl groups being optionally substituted.

[0194] In certain embodiments, spacer moieties Sp1and Sp2, if present, are independently selected from the group consisting of linear or branched C1-C50 alkylene groups, C2-C50 alkenylene groups, C2-C50 alkynylene groups, C3-C50 cycloalkylene groups, C5-C50 cycloalkenylene groups, C8-C50 cycloalkynylene groups, C7-C50 alkylarylene groups, C7-C50 arylalkylene groups, C8-C50 arylalkenylene groups and C9-C50 arylalkynylene groups, the alkylene groups, alkenylene groups, alkynylene groups, cycloalkylene groups, cycloalkenylene groups, cycloalkynylene groups, alkylarylene groups, arylalkylene groups, arylalkenylene groups and arylalkynylene groups being optionally substituted and optionally interrupted by one or more heteroatoms selected from the group of O, S and NR16, wherein R16is independently selected from the group consisting of hydrogen, Ci - C24 alkyl groups, C2 - C24 alkenyl groups, C2 - C24 alkynyl groups and C3 - C24 cycloalkyl groups, the alkyl groups, alkenyl groups, alkynyl groups and cycloalkyl groups being optionally substituted.

[0195] In certain embodiments, spacer moieties Sp1and Sp2, if present, are independently selected from the group consisting of linear or branched C1-C20 alkylene groups, C2-C20 alkenylene groups, C2-C20 alkynylene groups, C3-C20 cycloalkylene groups, C5-C20 cycloalkenylene groups, C8-C20 cycloalkynylene groups, C7-C20 alkylarylene groups, C7-C20 arylalkylene groups, C8-C20 arylalkenylene groups and C9-C20 arylalkynylene groups, the alkylene groups, alkenylene groups, alkynylene groups, cycloalkylene groups, cycloalkenylene groups, cycloalkynylene groups, alkylarylene groups, arylalkylene groups, arylalkenylene groups and arylalkynylene groups being optionally substituted and optionally interrupted by one or more heteroatoms selected from the group of O, S and NR16, wherein R16is independently selected from the group consisting of hydrogen, Ci - C24 alkyl groups, C2 - C24 alkenyl groups, C2 - C24 alkynyl groups and C3 - C24 cycloalkyl groups, the alkyl groups, alkenyl groups, alkynyl groups and cycloalkyl groups being optionally substituted.

[0196] In certain embodiments, the alkylene groups, alkenylene groups, alkynylene groups, cycloalkylene groups, cycloalkenylene groups, cycloalkynylene groups, alkylarylene groups, arylalkylene groups, arylalkenylene groups and arylalkynylene groups are unsubstituted and optionally interrupted by one or more heteroatoms selected from the group of O, S and NR16, e.g., O, wherein R16is independently selected from the group consisting of hydrogen and Ci - C4 alkyl groups, e.g., hydrogen or methyl.

[0197] In certain embodiments, spacer moieties Sp1and Sp2, if present, are independently selected from the group consisting of linear or branched C1-C20 alkylene groups, the alkylene groups being optionally substituted and optionally interrupted by one or more heteroatoms selected from the group of O, S and NR16, wherein R16is independently selected from the group consisting of hydrogen, Ci - C24 alkyl groups, C2 - C24 alkenyl groups, C2 - C24 alkynyl groups and C3 - C24 cycloalkyl groups, the alkyl groups, alkenyl groups, alkynyl groups and cycloalkyl groups being optionally substituted. In certain embodiments, the alkylene groups are unsubstituted and optionally interrupted by one or more heteroatoms selected from the group of O, S and NR16, e.g., O and / or S-S, wherein R16is independently selected from the group consisting of hydrogen and Ci - C4 alkyl groups, e.g., hydrogen or methyl.

[0198] In certain embodiments, Sp1and Sp2are -(CH2)r-, -(CH2CH2)r-, -(CH2CH2O)r-, -(OCH2CH2)r-, -(CH2CH2O)rCH2CH2-, -CH2CH2(OCH2CH2)r-, -(CH2CH2CH2O)r-, -(OCH2CH2CH2)r-, -(CH2CH2CH2O)rCH2CH2CH2-, or -CH2CH2CH2(OCH2CH2CH2)r-, wherein r is an integer in the range of 1 to 50, e.g., in the range of 1 to 40, in the range of 1 to 30, in the range of 1 to 20, or in the range of 1 to 15. In certain embodiments, n is 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10. In certain embodiments, n is 1, 2, 3, 4, 5, 6, 7 or 8. In certain embodiments, n is 1, 2, 3, 4, 5 or 6. In certain embodiments, n is 1, 2, 3 or 4.

[0199] In certain embodiments, linker L1is represented by -(W)k-(A)d-(B)e-(A)f-(C(O))g-wherein:d = 0 or 1, e.g., d = 1;e = an integer in the range 0- 10, e.g., e = 0, 1, 2, 3, 4, 5 or 6, e = an integer in the range 1 - 10, or e = 1, 2, 3 or 4;f = 0 or 1, e.g., f = 0;wherein d + e + f is at least 1, e.g., in the range 1 - 5, and / or wherein d + f is at least 1, e.g., d + f = 1.g = 0 or 1, e.g, g = 1;k = 0 or 1, e.g., k = 1;A is a sulfamide group according to structure (23);B is a -CH2-CH2-O- or a -O-CH2-CH2- moiety, or (B)eis a -(CH2-CH2-O)ei-CH2- CH2- or a - (CH2- CH2- O)ei- CH2- moiety, wherein el is defined the same way as e;W is -OC(O)-, -C(O)O-, -C(O)NH- -NHC(O)-, -OC(O)NH-, -NHC(O)O- -C(O)(CH2)mC(O)-, -C(O)(CH2)mC(O)NH- or -(4-Ph)CH2NHC(O)(CH2)mC(O)NH- e.g., wherein W is -OC(O)NH-, -C(0)(CH2)mC(0)NH- or -C(O)NH-, and wherein m is an integer in the range 0 - 10, e.g., m = 0, 1, 2, 3, 4, 5 or 6, or m = 2 or 3;optionally wherein L1is connected to Q via (W)k and to L2, L3, L4or D, e.g., to L2, via (C(O))g, e.g, via C(O).

[0200] In certain embodiments, the wavy lines in structure (23) represent the connection to the adjacent groups such as (W)k, (B)eand (C(O))g. In certain embodiments, A is according to structure (23), wherein a = 1 and R13= H or a Ci - C20 alkyl group, e.g., R13= H or methyl, or R13= H.

[0201] In certain embodiments, linker L1has the structure -(W)k-(A)d-(B)c-(A)f-(C(O))g-wherein:(a) k = 0; d = 1; g = 1; f = 0; B = -CH2-CH2-O-; e = 1, 2, 3 or 4, e.g., e = 2.(b) k = 1; W = -C(O)(CH2)mC(O)NH-; m = 2; d = 0; (B)e= -(CH2-CH2-O)ei-CH2- CH2-; f = 0; g = 1; el = 1, 2, 3 or 4, e.g, el = 1.(c) k = 1; W = -OC(O)NH-; d = 0; B = -CH2-CH2-O-; g = 1; f = 0; e = 1, 2, 3 or 4, e.g, e = 2.(d) k = 1; W = -C(O)(CH2)mC(O)NH-; m = 2; d = 0; (B)e= -(CH2-CH2-O)ei-CH2- CH2- f = 0; g = 1; el = 1, 2, 3 or 4, e.g., el = 4.(e) k = 1; W = -OC(O)NH-; d = 0; (B)e= -(CH2-CH2-O)ei-CH2-CH2-; g = 1; f = 0; el = 1, 2, 3 or 4, e.g., el = 4.(f) k = 1; W = -(4-Ph)CH2NHC(O)(CH2)mC(O)NH-, m = 3; d = 0; (B)e= -(CH2-CH2- O)ei-CH2-CH2-; g = 1; f = 0; el = 1, 2, 3 or 4, e.g, el = 4.(g) k = 0; d = 0; g = 1; f = 0; B = -CH2-CH2-O-; e = 1, 2, 3 or 4, e.g, e = 2.(h) k = 1; W = -C(O)NH-; d = 0; g = 1; f = 0; B = -CH2-CH2-O-; e = 1, 2, 3 or 4, e.g, e = 2.

[0202] Tn certain embodiments, linker L1comprises a branching nitrogen atom, which is located in the backbone between Q or Z and (L2)oand which comprises a further moiety D as substituent, which is optonally linked to the branching nitrogen atom via a linker. An example of a branching nitrogen atom is the nitrogen atom NR13in structure (23), wherein R13is connected to a second occurrence of D via a spacer moiety. Alternatively, a branching nitrogen atoms may be located within L1according to structure -(W)k-(A)d-(B)e-(A)f-(C(O))g-. In certain embodiments, L1is represented by -(W)k-(A)d-(B)e-(A)f-(C(O))g-N*[-(A)d-(B)e-(A)f-(C(O))g-]2, wherein A, B, W, d, e, f, g and k are as defined above and individually selected for each occurrence, and N* is the branching nitrogen atoms, to which two instances of -(A)d-(B)e-(A)f-(C(O))g- are connected. In certain embodiments, both (C(O))g- moi eties are connected to -(L2)0-(L3)p-(L4)q-D, wherein L2, L3, L4, o, p, q and D are as defined above and are each selected individually. In certain embodiments, each of L2, L3, L4, o, p, q and D are the same for both moieties connected to (C(O))g’.

[0203] In certain embodiments, linkers L1comprising a branching nitrogen atom have structure -( W)k-(A)d-(B)e-(A)f-(C(O))g-N* [-(A’ )d -(B ’ )e-(A’ )f’-(C(O))g-]2 wherein:(i) k = d = g = e’ = l; f = d’ = g’ = 0; W = -C(O)-; B = B’ = -CH2-CH2-O-; A is according to structure (23) with a = 0 and R13= H; e = 1, 2, 3 or 4, e.g., e = 2.(j) k = d = g = e’ = g’ = l; f = d’ = 0; W = -C(O)-; B = B’ = -CH2-CH2-O-; A is according to structure (23) with a = 0 and R13= H; e = 1, 2, 3 or 4, e.g., e = 2.Linker L2

[0204] Linker L2is either absent (o = 0) or present (o = 1). In certain embodiments, linker L2is present and 0 = 1. In certain embodiments, linker L2is a peptide spacer. In certain embodiments, the peptide spacer is defined by (NH-CR17-CO)n, wherein R17represents an amino acid side chain as known in the art. In certain embodiments, the amino acid may be a natural or a synthetic amino acid. In certain embodiments, the amino acid(s) are all in their L-configuration. In certain embodiments, n is an integer in the range of 1 - 5, e.g., in the range of 2 - 5. In certain embodiments, the peptide spacer comprises 1 - 5 amino acids. In certain embodiments, the peptide spacer is a dipeptide (n = 2), tripeptide (n = 3) or tetrapeptide (n = 4). In certain embodiments, the peptide spacer is a dipeptide. Although any peptide spacer may be used, in certain embodiments the peptide spacer is selected from Val-Cit, Vai-Ala, Val-Lys, Val-Arg, AcLys-Val-Cit, AcLys-Val-Ala, Glu-Val-Ala, Asp-Val-Ala, iGlu-Val-Ala, Glu-Val-Cit, Asp-Val-Cit, iGlu-Val-Cit, Phe-Cit, Phe-Ala, Phe-Lys, Phe-Arg, Ala-Lys, Leu-Cit, Ile-Cit, Trp-Cit, Ala-Ala-Asn, Ala-Asn, Gly-Gly-Phe-Gly and Lys. In certain embodiments, the peptide spacer is selected from Val-Cit, Val-Ala, Glu-Val-Ala, Val-Lys, Phe-Cit, Phe-Ala, Phe-Lys, and Ala-Ala-Asn. In certain embodiments, the peptide spacer is selected from Val-Cit, Vai-Ala, and Ala-Ala-Asn. In certain embodiments, the peptide spacer is selected from Val-Cit and Val-Ala. Herein, AcLys is acetyllysine and iGlu is isoglutamate. In certain embodiments, L2= Val-Cit. In certain embodiments, L2= Val-Ala.

[0205] R17represents the amino acid side chain, which is in certain embodiments selected from the side chains of alanine, cysteine, aspartic acid, glutamic acid, phenylalanine, glycine, histidine, isoleucine, lysine, acetyllysine, leucine, methionine, asparagine, pyrrolysine, proline, glutamine, arginine, serine, threonine, selenocysteine, valine, tryptophan, tyrosine and citrulline. In certain embodiments, the amino acid side chain is selected from those of Vai, Cit, Ala, Lys, Arg, AcLys, Phe, Leu, He, Trp, Glu, Asp and Asn. In certain embodiments, the amino acid side chain is selected from those of Vai, Cit, Ala, Glu and Lys. Alternatively worded, R17is in certain embodiments selected from CH3(Ala), CH2CH(CH3)2(Leu), CH2CH2CH2NHC(O)NH2 (Cit), CH2CH2CH2CH2NH2(Lys), CH2CH2CH2NHC(O)CH3(AcLys), CH2CH2CH2NHC(=NH)NH2(Arg), CH2Ph (Phe), CH(CH3)2(Vai), CH(CH3)CH2CH3(He), CH2C(O)NH2(Asn), CH2CH2C(O)OH (Glu), CH2C(O)OH (Asp) and CH2(lH-indol-3-yl) (Trp). In certain embodiments, R17is selected from CH3(Ala), CH2CH2CH2NHC(O)NH2(Cit), CH2CH2CH2CH2NH2(Lys), CH2CH2C(O)OH (Glu) and CH(CH3)2(Vai). In certain embodiments, R17is CH3(Ala), CH2CH2CH2NHC(O)NH2(Cit), CH2CH2CH2CH2NH2(Lys), or CH(CH3)2(Vai).

[0206] In certain embodiments, the peptide spacer is represented by general structure (L3):wherein R17is as defined above. In certain embodiments, R17is CH3(Vai) or CH2CH2CH2NHC(O)NH2(Cit). The wavy lines in general structure (L3) indicate the connection to (L1),, and (L3)p, e.g., L2according to structure (L3) is connected to (L1),, via NH and to (L3)pvia C(O).Linker L3

[0207] Linker L3is either absent (p = 0) or present (p = 1). In certain embodiments, linker L3is present and p = 1. In certain embodiments, linker L3is a self-cleavable spacer, also referred toas self-immolative spacer. Tn certain embodiments, L3is a para-aminobenzyl oxy carbonyl (PABC) derivative. In certain embodiments, L3is a PABC derivative according to structure (L4):wherein the wavy lines indicate the connection to Q or Z, L1or L2, and to L4or D. In certain embodiments, the PABC derivative is connected via NH to Q, Z, L1or L2, e.g, to L2, and via O to L4or D.

[0208] A is a 5- or 6-membered aromatic or heteroaromatic ring, e.g., a 6-membered aromatic or heteroaromatic ring. Suitable 5-membered rings are oxazole, thiazole and furan. Suitable 6-membered rings are phenyl and pyridyl. In certain embodiments, A is 1,4-phenyl, 2,5-pyridyl or 3,6-pyridyl. In certain embodiments, A is 1,4-phenyl.

[0209] R21is selected from H, R26, C(O)OH and C(O)R26, wherein R26is Ci - C24 (hetero)alkyl groups, C3 - C10 (hetero)cycloalkyl groups, C2 - C10 (hetero)aryl groups, C3 - C10 alkyl(hetero)aryl groups and C3 - C10 (hetero)arylalkyl groups, which are optionally substituted and optionally interrupted by one or more heteroatoms selected from O, S and NR28wherein R28is independently selected from the group consisting of hydrogen and Ci - C4 alkyl groups. In certain embodiments, R26is C3 - C10 (hetero)cycloalkyl or polyalkylene glycol. In certain embodiments, the polyalkylene glycol is a polyethylene glycol or a polypropylene glycol. In certain embodiments, the polyalkylene glycol is -(CH2CH2O)SH or -(CH₂CH₂CH₂O)sH. In certain embodiments, the polyalkylene glycol is a polyethylene glycol, e.g, -(CH2CH2O)SH, wherein s is an integer in the range 1 - 10, e.g., s is an integer in the range 1 - 5, or s = 1, 2, 3 or 4. In certain embodiments, R21is H or C(O)R26, wherein R26= 4-methyl-piperazine or morpholine. In certain embodiments, R21is H.Linker L4

[0210] Linker L4is either absent (q = 0) or present (q = 1). In certain embodiments, linker L4is present and q = 1. In certain embodiments, linker L4is selected from:an aminoalkanoic acid spacer according to the structure - NR22-(Cz-alkylene)-C(O)-, wherein z is an integer in the range 1 - 20 and R22is H or Ci - C4 alkyl;an ethyleneglycol spacer according to the structure -NR22-(CH2-CH2-O)e6-(CH2)e7- C(O)-, wherein e6 is an integer in the range 1 - 10, el is an integer in the range 1 -3 and R22is H or Ci - C4 alky; anda diamine spacer according to the structure - NR22-(Cz-alkylene)-NR22-(C(O))h-, wherein h is 0 or 1, z is an integer in the range 1 - 20 and R22is H or Ci - C4 alkyl.

[0211] Linker L4may be an aminoalkanoic acid spacer, z.e., -NR22-(Cz-alkylene)-C(O)-, wherein z is an integer in the range 1 to 20, e.g., 1 - 10 or 1 - 6. In certain embodiments, the aminoalkanoic acid spacer is connected to L3via the nitrogen atom and to D via the carbonyl moiety. In certain embodiments, linker L4is selected from 6-aminohexanoic acid (Ahx, z = 5), 0-alanine (z = 2) and glycine (Gly, z = 1). In certain embodiments, linker L4is selected from 6-aminohexanoic acid or glycine. In certain embodiments, linker L4is 6-aminohexanoic acid. In certain embodiments, linker L4is glycine. In certain embodiments, R22is H or Ci - C4 alkyl. In certain embodiments, R22is H or methyl. In certain embodiments, R22is H.

[0212] Alternatively, linker L4may be an ethyleneglycol spacer according to the structure -NR22(CH2CH2O)e6 (CH2)e7 -(C(O)-, wherein e6 is an integer in the range 1 - 10, e.g., e6 is in the range 2-6, and e7 is an integer in the range 1 -3, e.g., el is 2. In certain embodiments, R22is H or Ci - C4 alkyl. In certain embodiments, R22is H or methyl. In certain embodiments, R22is H.

[0213] In certain embodiments, linker L4is a diamine spacer according to the structure - NR22-(Cz-alkylene)-NR22-(C(O))h-, wherein h is 0 or 1, z is an integer in the range 1 - 20, e.g., z is an integer in the range 2 - 6, z = 2 or 5, or z = 2. In certain embodiments, R22is H or Ci - C4 alkyl. In certain embodiments, R22is H or Ci - C4 alkyl. In certain embodiments, R22is H or methyl. In certain embodiments, R22is methyl. In certain embodiments, h is 1, in which case linker L4is especially suited for conjugation via a phenolic hydroxyl group present on payload D.Payload D

[0214] D represents the target molecule D that is or is to be connected to the antibody, which is also referred to in the art as the payload. In certain embodiments, D is selected from any of the pharmaceutically active substances described above. In certain embodiments, D is selected from the group consisting of taxanes, anthracyclines, camptothecins, epothilones, mytomycins, combretastatins, vinca alkaloids, tubulysins, enediynes (e.g., calicheamicins), amanitins, duocarmycins, maytansinoids, amatoxins, dolastatins, auristatins, eribulins, BCL-XL inhibitors, hemiasterlins, KSP inhibitors, radioisotopes, TLR agonists, indolinobenzodiazepine dimers orpyrrolobenzodiazepine dimers (PBDs), and analogues or prodrugs thereof. Tn certain embodiments, D is defined as a pharmaceutically active substance, such as an anti-cancer agent, e.g., a cytotoxin. In certain embodiments, D is selected from the group consisting of anthracyclines, camptothecins, maytansinoids, enediynes, amanitins, auristatins and pyrrolobenzodiazepine dimers. In certain embodiments, D is selected from the group consisting of enediynes, auristatins and camptothecins. In certain embodiments, D is an enediyne. In certain embodiments, D is an auristatin. In certain embodiments, D is a camptothecin.

[0215] In certain embodiments, the enediyne is selected from the group consisting of calicheamicins, esperamicins, shishijimicins and namenamicins. In certain embodiments, the enediyne is calicheamicin. In certain embodiments, the auristatin is selected from the group consisting of MMAD, MMAE, MMAF or PF-06380101. In certain embodiments, the auristatin is MMAE or PF-06380101. In certain embodiments, the auristatin is MMAE. In certain embodiments, the camptothecin is selected from the group consisting of topotecan, silatecan, cositecan, exatecan, exatecan-S, DXd, SN-38, lurtotecan, gimatecan, belotecan, rubitecan, AMDCPT and G-AMDCPT. In certain embodiments, the camptothecin is exatecan or DXd. In certain embodiments, the camptothecin is exatecan.[00216J In certain embodiments, D is selected from the group consisting of calicheamicin, MMAE, PF-06380101, exatecan and DXd. In certain embodiments, D is selected from the group consisting of calicheamicin, MMAE, and exatecan. In certain embodiments, D is selected from the group consisting of MMAE and exatecan. In certain embodiments, D is MMAE. In certain embodiments, D is exatecan.

[0217] The compound according to general structure (2) may comprise more than one moiety D. When more than one cytotoxin D is present, the cytotoxins D may be the same or different. In certain embodiments, the compound according to general structure (2) comprises 1 or 2 occurrences of D, e.g., 2 occurrence of D. In certain embodiments, the second occurrence of D is present within linker L, which may comprise a branching moiety, typically a branching nitrogen atom, that is connected to the second occurrence of D. In certain embodiments, both occurrences of D are connected to the branching moiety via the same linker. Likewise, the ADC according to structure (1) may comprise more than one moiety D per connecting group Z.Exemplary A DCs

[0218] Exemplary ADCs of the present disclosure include compounds (I) - (III) and (X) -(XIII). In certain embodiments, the ADC is selected from (Xa), (Xlb), (Xllg), (Xllh) and (XHIe).In certain embodiments, the ADC is selected from (XI) and (XIII). In certain embodiments, the ADC is (Xlb) or (XHIe). In certain embodiments, the ADC is according to (XIII). In certain embodiments, the ADC is according to (XHIe). The structures of these ADCs are defined here below.

[0219] ADC (I) has the following structure:AB-[(L6)-{Z-(L1)-(L2)-(L3)-(L4)q-D}x]y(1)wherein:AB, L6, Z, D, x and y are as defined above;L1is a linker represented by -(A)d-(B)e-(A)f-(C(O))g- as defined above;L2is Val-Cit or Vai-Ala;L3is the PABC derivative according to structure (L4);L4is -N-(Cz-alkylene)-C(O)- or -NR22-(Cz-alkylene)-NR22-, wherein z and R22are as defined above;q = 0 or 1.

[0220] In certain embodiments of ADC (I), for L1, d = 1, A is according to structure (23), e.g., wherein a = 1 and R13= H, e = 2, f = 0 and g = 1. In certain embodiments of ADC (I), L2= Val-Cit. In certain embodiments of ADC (I), for L3, R21= H. In certain embodiments of ADC (I), in case q = 1, then z = 1 or 5.

[0221] In certain embodiments, the ADC according to structure (I) comprises a payload D selected from the group consisting of calicheamicin, MMAE, PF-06380101, exatecan and DXd. In certain embodiments, payload D is selected from the group consisting of calicheamicin, MMAE, and exatecan. In certain embodiments, payload D is exatecan.

[0222] ADC (II) has the following structure:AB-[(L6)-{Z-(L1)-(L2)-(L3)-D}x]y(11)wherein:AB, L6, Z, D, x and y are as defined above;L1is a linker represented by -(A)-(B)e-(C(O))-, as defined above;L2is Val-Cit or Vai-Ala;L3is the PABC derivative according to structure (L4), wherein R21= H.

[0223] In certain embodiments of ADC (II), for L1, e = 2, A is according to structure (23), e.g., wherein a = 1 and R13= H. In certain embodiments of ADC (II), L2= Val-Cit.

[0224] In certain embodiments, the ADC according to structure (II) comprises a payload D selected from the group consisting of calicheamicin, MMAE, PF-06380101, exatecan and DXd. In certain embodiments, payload D is selected from the group consisting of calicheamicin, MMAE, and exatecan. In certain embodiments, payload D is exatecan.

[0225] ADC (III) has the following structure:AB-[(L6)-{Z-(L1)-(L2)-(L3)-(L4)-D}x]y(HI)wherein:AB, L6, Z, D, x and y are as defined above;L1is a linker represented by -(A)-(B)e-(C(O))-, as defined above;L2is Val-Cit or Vai-Ala;L3is the PABC derivative according to structure (L4), wherein R21= H;L4is -NR22-(Cz-alkylene)-NR22-, wherein R22is as defined above and z is an integer in the range 1 to 6.

[0226] In certain embodiments of ADC (III), for L1, e = 2, A is according to structure (23), e.g., wherein a = 1 and R13= H. In certain embodiments of ADC (III), L2= Val-Cit. In certain embodiments of ADC (III), z = 2.

[0227] In certain embodiments, the ADC according to structure (III) comprises a payload D selected from the group consisting of calicheamicin, MMAE, PF-06380101, exatecan and DXd. In certain embodiments, payload D is selected from the group consisting of calicheamicin, MMAE, and exatecan. In certain embodiments, payload D is exatecan.

[0228] ADC (X) has a linker-payload moiety according to the following structure:wherein:the wavy line indicates the connection to Z;L2, o and D are as defined above.

[0229] L2may be present or absent. In certain embodiments, L2is present and o = 1. For ADC embodiment (Xa), L2is according to structure (L3) and R17is CH3. For ADC embodiment (Xb), L2is according to structure (L3) and R17is CH2CH2CH2NHC(O)NH2. In certain embodiments, ADC (X) has structure (Xa).

[0230] In certain embodiments, the ADC according to structure (X) comprises a payload D selected from the group consisting of calicheamicin, MMAE, PF-06380101, exatecan and DXd. In certain embodiments, payload D is selected from the group consisting of calicheamicin, MMAE, and exatecan. In certain embodiments, payload D is exatecan.

[0231] ADC (XI) has a linker-payload moiety according to the following structure:wherein:the wavy line indicates the connection to Z;L2, 0 and D are as defined above.

[0232] L2may be present or absent. In certain embodiments, L2is present and o = 1. For ADC embodiment (Xia), L2is according to structure (L3) and R17is CH3. For ADC embodiment (Xlb), L2is according to structure (L3) and R17is CH2CH2CH2NHC(O)NH2. In certain embodiments, ADC (XI) has structure (Xlb).

[0233] In certain embodiments, the ADC according to structure (XT) comprises a payload D selected from the group consisting of calicheamicin, MMAE, PF-06380101, exatecan and DXd. In certain embodiments, payload D is selected from the group consisting of calicheamicin, MMAE, and exatecan. In certain embodiments, payload D is calicheamicin.

[0234] ADC (XII) has a linker-payload moiety according to the following structure:wherein:the wavy line indicates the connection to Z;L2, L4, o, q and D are as defined above.

[0235] L2may be present or absent. In certain embodiments, L2is present and o = 1. For ADC embodiment (Xlla), L2is according to structure (L3) and R17is CH3. For ADC embodiment (Xllb), L2is according to structure (L3) and R17is CH2CH2CH2NHC(O)NH2. In certain embodiments, R17= CH2CH2CH2NHC(O)NH2.

[0236] L4may be present or absent. For ADC embodiment (XIIc), q = 0 and L4is absent. For ADC embodiment (XTId), q = 1 and L4is a diamine spacer according to the structure -NR22-(CZ-alkylene)-NR22-, wherein z is an integer in the range 1 - 20, and R22is H or Ci - C4 alkyl.

[0237] For ADC embodiment (Xlle), L2is according to structure (L3), R17is CH3, q = 0 and L4is absent. For ADC embodiment (Xllf), L2is according to structure (L3), R17is CH3, q = 1 and L4is a diamine spacer according to the structure -NR22-(Cz-alkylene)-NR22-, wherein z is an integer in the range 1 - 10, e.g., wherein z = 2, and R22is H or Ci - C4 alkyl.

[0238] For ADC embodiment (Xllg), L2is according to structure (L3), R17is CH2CH2CH2NHC(O)NH2, q = 0 and L4is absent. For ADC embodiment (Xllh), L2is according to structure (L3), R17is CH2CH2CH2NHC(O)NH2, q = 1 and L4is a diamine spacer according tothe structure -NR22-(Cz-alkylene)-NR22-, wherein z is an integer in the range 1 - 20, and R22is H or Ci - C4 alkyl.[002391 In certain embodiments of ADC (XII), z is an integer in the range of 1 - 10, e.g., z = 2 - 6, or z = 2. In certain embodiments of ADC (XII), R22is H or CH3.

[0240] In certain embodiments, ADC (XII) has structure (Xllg) or structure (Xllh).

[0241] In certain embodiments, the ADC according to structure (XII) comprises a payload D selected from the group consisting of calicheamicin, MMAE, PF-06380101, exatecan and DXd. In certain embodiments, payload D is selected from the group consisting of calicheamicin, MMAE, and exatecan. In certain embodiments, payload D is MMAE. In certain embodiments, payload D is exatecan.

[0242] ADC (XIII) has a linker-payload moiety according to the following structure:wherein:the wavy line indicates the connection to Z;L2, L4, o, q and D are as defined above.

[0243] L2may be present or absent. In certain embodiments, L2is present and o = 1. For ADC embodiment (XHIa), L2is according to structure (L3) and R17is CH3. For ADC embodiment (XHIb), L2is according to structure (L3) and R17is CH^CFkCIENHCIOjNFh. In certain embodiments, R17= CH3.

[0244] L4may be present or absent. For ADC embodiment (XIIIc), q = 0 and L4is absent. For ADC embodiment (XHId), q = 1 and L4is a diamine spacer according to the structure -NR22-(CZ-alkylene)-NR22-, wherein z is an integer in the range 1 - 20, and R22is H or Ci - C4 alkyl.

[0245] For ADC embodiment (XIITe), L2is according to structure (L3), R17is CH3, q = 0 and L4is absent. For ADC embodiment (XHIf), L2is according to structure (L3), R17is CH3, q = 1 and L4is a diamine spacer according to the structure -NR22-(Cz-alkylene)-NR22-, wherein z is an integer in the range 1 - 10, e.g., wherein z = 2, and R22is H or Ci - C4 alkyl.

[0246] For ADC embodiment (XHIg), L2is according to structure (L3), R17is CH2CH2CH2NHC(O)NH2, q = 0 and L4is absent. For ADC embodiment (XHIh), L2is according to structure (L3), R17is CH2CH2CH2NHC(O)NH2, q = 1 and L4is a diamine spacer according to the structure -NR22-(Cz-alkylene)-NR22-, wherein z is an integer in the range 1 - 20, and R22is H or Ci - C4 alkyl.

[0247] In certain embodiments of ADC (XIII), z is an integer in the range of 1 - 10, e.g., z is an integer in the range of 2 - 6, or z = 2. In certain embodiments of ADC (XIII), R22is H or CH3.

[0248] In certain embodiments, ADC (XIII) has structure (XHIe).

[0249] In certain embodiments, the ADC according to structure (XIII) comprises a payload D selected from the group consisting of calicheamicin, MMAE, PF-06380101, exatecan and DXd. In certain embodiments, payload D is selected from the group consisting of calicheamicin, MMAE, and exatecan. In certain embodiments, payload D is MMAE. In certain embodiments, payload D is exatecan.

[0250] In certain embodiments, the ADCs are conjugated through the glycan, z.e., b = 1. In certain embodiments, the ADCs are conjugated through a trimmed glycan, z.e., j = 0. In certain embodiments, S = GalNAc and w’ = 0. In certain embodiments, connecting group Z is formed by an azide-alkyne cycloaddition, e.g, connecting group Z = (Z39), wherein ring Z = (Za) and V = CH2. In certain embodiments, connecting group Z = (Z39), wherein ring Z = (Za) and V = CH2. In certain embodiments, x = 1. In certain embodiments, y = 2. In certain embodiments, x = 1 and y = 2.

[0251] In certain embodiments, the ADC is an ADC of general structure (1), wherein L-D (the linker-payload moiety) is as in ADC embodiment (Xllg), wherein payload D is MMAE; connecting group Z is formed by a reaction (e.g, an azide-alkyne cycloaddition) between compatible click probe moi eties, e.g, connecting group Z = (Z39), wherein ring Z = (Za) and V = CH2 or connecting group Z = (Z42), wherein all R15, all R18, and R19are each hydrogen and 1 is 1; b is 1 and L6is as defined above; AB is an antibody that specifically binds to LY6K (e.g., human LY6K); x is 1; and y is 2. In certain embodiments, L6is connected to an amino acid at the conservedN-glycosylation site at the asparagine residue at or close to position 297 of the heavy chain. In certain embodiments, L6is connected to an amino acid at the conserved N-glycosylation site at the asparagine residue at position 297 of the heavy chain. In certain embodiments, the ADC is as shown in FIG. 2A.

[0252] In certain embodiments, the ADC is an ADC of general structure (1), wherein L-D (the linker-payload moiety) is as in ADC embodiment (Xllg), wherein payload D is MMAE; connecting group Z = (Z39), wherein ring Z = (Za) and V = CH2; b is 1 and L6is as defined above; AB is an antibody that specifically binds to LY6K (e.g., human LY6K); x is 1; and y is 2.

[0253] In certain embodiments, the ADC is an ADC of general structure (1), wherein L-D (the linker-payload moiety) is as in ADC embodiment (Xllg), wherein payload D is MMAE; connecting group Z = (Z42), wherein all R13, all R18, and R19are each hydrogen and 1 is 1; b is 1 and L6is as defined above; AB is an antibody that specifically binds to LY6K (e.g, human LY6K); x is 1; and y is 2.

[0254] In certain embodiments, the ADC is an ADC of general structure (1), wherein L-D (the linker-payload moiety) is as in ADC embodiment (Xllg), wherein payload D is MMAE; connecting group Z is formed by an azide-alkyne cycloaddition, e.g., connecting group Z = (Z39), wherein ring Z = (Za) and V = CH2 or connecting group Z = (Z42), wherein all R13, all R18, and R19are each hydrogen and 1 is 1; b is 1 and L6is -GlcNAc(Fuc)w-(G)j-S-(L7)W’-, wherein j and w’ are each 0, w is 0 or 1, and S is GalNAc; AB is an antibody that specifically binds to LY6K (e.g., human LY6K); x is 1; and y is 2. In certain embodiments, the antibody comprises: a VH comprising the CDRH1, CDRH2, and CDRH3 amino acid sequences of the VH amino acid sequence set forth in any one of SEQ ID NOs: 1-34; and a VL comprising the CDRL1, CDRL2, and CDRL3 amino acid sequences of the VL amino acid sequence set forth in any one of SEQ ID NOs: 35-68. In certain embodiments, the VH amino acid sequence and VL amino acid sequence are as set forth in: SEQ ID NOs: 2 and 36 or 16 and 50, respectively. In certain embodiments, the antibody comprises the CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in: SEQ ID NOs: 70, 91, 125, 159, WAS, and SEQ ID NO: 199 or SEQ ID NOs: 74, 105, 138, 170, 185, and 211, respectively. In certain embodiments, the antibody comprises the VH amino acid sequence and VL amino acid sequence set forth in SEQ ID NOs: 2 and 36 or 16 and 50, respectively. In certain embodiments, L6is connected to an amino acid at the conserved N-glycosylation site at the asparagine residue at or close to position 297 of the heavychain. In certain embodiments, L6is connected to an amino acid at the conserved N-glycosylation site at the asparagine residue at position 297 of the heavy chain. In certain embodiments, the ADC is as shown in FIG. 2B or FIG. 2C. In certain embodiments, w is 1 (FIG. 2B). In certain embodiments, w is 1 (FIG. 2C).

[0255] In certain embodiments, the ADC is an ADC of general structure (1), wherein L-D (the linker-payload moiety) is as in ADC embodiment (Xllg), wherein payload D is MMAE; connecting group Z = (Z39), wherein ring Z = (Za) and V = CH2; b is 1 and L6is -GlcNAc(Fuc)w-(G)j-S-(L7)W- wherein j and w’ are each 0, w is 0 or 1, and S is GalNAc; AB is an antibody that specifically binds to LY6K (e.g., human LY6K); x is 1; and y is 2. In certain embodiments, the antibody comprises: a VH comprising the CDRH1, CDRH2, and CDRH3 amino acid sequences of the VH amino acid sequence set forth in any one of SEQ ID NOs: 1-34; and a VL comprising the CDRL1, CDRL2, and CDRL3 amino acid sequences of the VL amino acid sequence set forth in any one of SEQ ID NOs: 35-68. In certain embodiments, the VH amino acid sequence and VL amino acid sequence are as set forth in: SEQ ID NOs: 2 and 36 or 16 and 50, respectively. In certain embodiments, the antibody comprises the CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, andCDRL3 amino acid sequences set forth in: SEQ ID NOs: 70, 91, 125, 159, WAS, and SEQ ID NO: 199 or SEQ ID NOs: 74, 105, 138, 170, 185, and 211, respectively. In certain embodiments, the antibody comprises the VH amino acid sequence and VL amino acid sequence set forth in SEQ ID NOs: 2 and 36 or 16 and 50, respectively. In certain embodiments, L6is connected to an amino acid at the conserved N-glycosylation site at the asparagine residue at or close to position 297 of the heavy chain. In certain embodiments, L6is connected to an amino acid at the conserved N-glycosylation site at the asparagine residue at position 297 of the heavy chain. In certain embodiments, w is 1 (FIG. 2B) In certain embodiments, w is 1 (FIG. 2C)

[0256] In certain embodiments, the ADC is an ADC of general structure (1), wherein L-D (the linker-payload moiety) is as in ADC embodiment (Xllg), wherein payload D is MMAE; connecting group Z = (Z42), wherein all R15, all R18, and R19are each hydrogen and 1 is 1; b is 1 and L6is -GlcNAc(Fuc)w-(G)j-S-(L7)w-, wherein j and w’ are each 0, w is 0 or 1, and S is GalNAc; AB is an antibody that specifically binds to LY6K (e.g., human LY6K); x is 1; and y is 2. In certain embodiments, the antibody comprises: a VH comprising the CDRH1, CDRH2, and CDRH3 amino acid sequences of the VH amino acid sequence set forth in any one of SEQ ID NOs: 1-34; and a VL comprising the CDRL1, CDRL2, and CDRL3 amino acid sequences of theVL amino acid sequence set forth in any one of SEQ ID NOs: 35-68. In certain embodiments, the VH amino acid sequence and VL amino acid sequence are as set forth in: SEQ ID NOs: 2 and 36 or 16 and 50, respectively. In certain embodiments, the antibody comprises the CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in: SEQ ID NOs: 70, 91, 125, 159, WAS, and SEQ ID NO: 199 or SEQ ID NOs: 74, 105, 138, 170, 185, and 211, respectively. In certain embodiments, the antibody comprises the VH amino acid sequence and VL amino acid sequence set forth in SEQ ID NOs: 2 and 36 or 16 and 50, respectively. In certain embodiments, L6is connected to an amino acid at the conserved N-glycosylation site at the asparagine residue at or close to position 297 of the heavy chain. In certain embodiments, L6is connected to an amino acid at the conserved N-glycosylation site at the asparagine residue at position 297 of the heavy chain. In certain embodiments, w is 1 (FIG.2B). In certain embodiments, w is 1 (FIG. 2C).

[0257] In certain embodiments, the ADC is an ADC of general structure (1), wherein L-D (the linker-payload moiety) is as in ADC embodiment (XIHe), wherein payload D is exatecan; connecting group Z is formed by a reaction (e.g., an azi de-alkyne cycloaddition) between compatible click probe moi eties, e.g., connecting group Z = (Z39), wherein ring Z = (Za) and V = CH2 or connecting group Z = (Z42), wherein all R13, all R18, and R19are each hydrogen and 1 is 1; b is 1 and L6is as defined above; AB is an antibody that specifically binds to LY6K (e.g., human LY6K); x is 1; and y is 2. In certain embodiments, L6is connected to an amino acid at the conserved N-glycosylation site at the asparagine residue at or close to position 297 of the heavy chain. In certain embodiments, L6is connected to an amino acid at the conserved N-glycosylation site at the asparagine residue at position 297 of the heavy chain. In certain embodiments, the ADC is as shown in FIG. 2D.

[0258] In certain embodiments, the ADC is an ADC of general structure (1), wherein L-D (the linker-payload moiety) is as in ADC embodiment (XIHe), wherein payload D is exatecan; connecting group Z = (Z39), wherein ring Z = (Za) and V = CH2; b is 1 and L6is as defined above; AB is an antibody that specifically binds to LY6K (e.g., human LY6K); x is 1; and y is 2.

[0259] In certain embodiments, the ADC is an ADC of general structure (1), wherein L-D (the linker-payload moiety) is as in ADC embodiment (XIHe), wherein payload D is exatecan; connecting group Z = (Z42), wherein all R13, all R18, and R19are each hydrogen and 1 is 1; b is 1and L6is as defined above; AB is an antibody that specifically binds to LY6K (e.g, human LY6K); x is 1; and y is 2.

[0260] In certain embodiments, the ADC is an ADC of general structure (1), wherein L-D (the linker-payload moiety) is as in ADC embodiment (XHIe), wherein payload D is exatecan; connecting group Z is formed by an azide-alkyne cycloaddition, e.g., connecting group Z = (Z39), wherein ring Z = (Za) and V = CH2 or connecting group Z = (Z42), wherein all R13, all R18, and R19are each hydrogen and 1 is 1; b is 1 and L6is -GlcNAc(Fuc)w-(G)j-S-(L7)W’-, wherein j and w’ are each 0, w is 0 or 1, and S is GalNAc; AB is an antibody that specifically binds to LY6K (e.g, human LY6K); x is 1; and y is 2. In certain embodiments, the antibody comprises: a VH comprising the CDRH1, CDRH2, and CDRH3 amino acid sequences of the VH amino acid sequence set forth in any one of SEQ ID NOs: 1-34; and a VL comprising the CDRL1, CDRL2, and CDRL3 amino acid sequences of the VL amino acid sequence set forth in any one of SEQ ID NOs: 35-68. In certain embodiments, the VH amino acid sequence and VL amino acid sequence are as set forth in: SEQ ID NOs: 2 and 36 or 16 and 50, respectively. In certain embodiments, the antibody comprises the CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in: SEQ ID NOs: 70, 91, 125, 159, WAS, and SEQ ID NO: 199 or SEQ ID NOs: 74, 105, 138, 170, 185, and 211, respectively. In certain embodiments, the antibody comprises the VH amino acid sequence and VL amino acid sequence set forth in SEQ ID NOs: 2 and 36 or 16 and 50, respectively. In certain embodiments, L6is connected to an amino acid at the conserved N-glycosylation site at the asparagine residue at or close to position 297 of the heavy chain. In certain embodiments, L6is connected to an amino acid at the conserved N-glycosylation site at the asparagine residue at position 297 of the heavy chain. In certain embodiments, the ADC is as shown in FIG. 2E or FIG. 2F. In certain embodiments, w is 1 (FIG. 2E). In certain embodiments, w is 1 (FIG.2F)

[0261] In certain embodiments, the ADC is an ADC of general structure (1), wherein L-D (the linker-payload moiety) is as in ADC embodiment (XHIe), wherein payload D is exatecan; connecting group Z = (Z39), wherein ring Z = (Za) and V = CH2; b is 1 and L6is -GlcNAc(Fuc)w-(G)j-S-(L7)w - wherein j and w’ are each 0, w is 0 or 1, and S is GalNAc; AB is an antibody that specifically binds to LY6K (e.g., human LY6K); x is 1; and y is 2. In certain embodiments, the antibody comprises: a VH comprising the CDRH1, CDRH2, and CDRH3 amino acid sequences of the VH amino acid sequence set forth in any one of SEQ ID NOs: 1-34; and a VL comprisingthe CDRL1, CDRL2, and CDRL3 amino acid sequences of the VL amino acid sequence set forth in any one of SEQ ID NOs: 35-68. In certain embodiments, the VH amino acid sequence and VL amino acid sequence are as set forth in: SEQ ID NOs: 2 and 36 or 16 and 50, respectively. In certain embodiments, the antibody comprises the CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, andCDRL3 amino acid sequences set forth in: SEQ ID NOs: 70, 91, 125, 159, WAS, and SEQ ID NO: 199 or SEQ ID NOs: 74, 105, 138, 170, 185, and 211, respectively. In certain embodiments, the antibody comprises the VH amino acid sequence and VL amino acid sequence set forth in SEQ ID NOs: 2 and 36 or 16 and 50, respectively. In certain embodiments, L6is connected to an amino acid at the conserved N-glycosylation site at the asparagine residue at or close to position 297 of the heavy chain. In certain embodiments, L6is connected to an amino acid at the conserved N-glycosylation site at the asparagine residue at position 297 of the heavy chain. In certain embodiments, w is 1 (FIG. 2E). In certain embodiments, w is 1 (FIG. 2F).

[0262] In certain embodiments, the ADC is an ADC of general structure (1), wherein L-D (the linker-payload moiety) is as in ADC embodiment (XIHe), wherein payload D is exatecan; connecting group Z = (Z42), wherein all R15, all R18, and R19are each hydrogen and 1 is 1; b is 1 and L6is -GlcNAc(Fuc)w-(G)j-S-(L7)w-, wherein j and w’ are each 0, w is 0 or 1, and S is GalNAc; AB is an antibody that specifically binds to LY6K (e.g., human LY6K); x is 1; and y is 2. In certain embodiments, the antibody comprises: a VH comprising the CDRH1, CDRH2, and CDRH3 amino acid sequences of the VH amino acid sequence set forth in any one of SEQ ID NOs: 1-34; and a VL comprising the CDRL1, CDRL2, and CDRL3 amino acid sequences of the VL amino acid sequence set forth in any one of SEQ ID NOs: 35-68. In certain embodiments, the VH amino acid sequence and VL amino acid sequence are as set forth in: SEQ ID NOs: 2 and 36 or 16 and 50, respectively. In certain embodiments, the antibody comprises the CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in: SEQ ID NOs: 70, 91, 125, 159, WAS, and SEQ ID NO: 199 or SEQ ID NOs: 74, 105, 138, 170, 185, and 211, respectively. In certain embodiments, the antibody comprises the VH amino acid sequence and VL amino acid sequence set forth in SEQ ID NOs: 2 and 36 or 16 and 50, respectively. In certain embodiments, L6is connected to an amino acid at the conserved N-glycosylation site at the asparagine residue at or close to position 297 of the heavy chain. In certain embodiments, L6is connected to an amino acid at the conserved N-glycosylation site at the asparagine residue atposition 297 of the heavy chain. In certain embodiments, w is 1 (FIG.2E). In certain embodiments, w is 1 (FIG. 2F).Compound according to general structure (2)

[0263] The compound has general structure (2):Q-L-D(2)wherein:Q is a reactive moiety;L is a linker that links Z to D;D is selected from the group consisting of taxanes, anthracyclines, camptothecins, epothilones, mytomycins, combretastatins, vinca alkaloids, tubulysins, enediynes (e.g., calicheamicins), amanitins, duocarmycins, maytansinoids, amatoxins, dolastatins, auristatins, eribulins, BCL-XL inhibitors, hemiasterlins, KSP inhibitors, radioisotopes, TLR agonists, indolinobenzodi azepine dimers or pyrrolobenzodiazepine dimers (PBDs), and analogues or prodrugs thereof;

[0264] The compound of general structure (2) may also be referred to as a “linker-drug construct”, for comprising linker L and payload D of the final conjugate. Compounds according to general structure (2) can be prepared by the skilled person using standard organic synthesis techniques. Linker L and payload D are defined above in the context of the conjugate according to structure (1).Reactive moiety 0

[0265] The compound according to general structure (2) comprises a reactive moiety Q. In the context of the present disclosure, the term “reactive moiety” may refer to a chemical moiety that comprises a reactive group, but also to a reactive group itself. For example, a cyclooctynyl group is a reactive group comprising a reactive group, namely a C-C triple bond. Similarly, an N-maleimidyl group is a reactive group, comprising a C-C double bond as a reactive group. However, a reactive group, for example an azido reactive group, a thiol reactive group or an alkynyl reactive group, may herein also be referred to as a reactive moiety.

[0266] Q serves as chemical handle for the connection to S(F)X. In other words, Q is reactive towards and complementary to F. Herein, a reactive group is denoted as “complementary” to areactive group when said reactive group reacts with said reactive group selectively, optionally in the presence of other functional groups. Complementary reactive and functional groups are known to a person skilled in the art, and are described in more detail below. As such, the compound according to general structure (2) is conveniently used in a conjugation reaction, wherein a chemical reaction between Q and F takes place, thereby forming an ADC comprising a covalent connection between payload D and the antibody.

[0267] The exact nature of Q, and F, depends on the type of conjugation reaction that is employed. The skilled person will be able to select the appropriate combination of Q and F. In certain embodiments, Q, and thus also F, is reactive in a cycloaddition or a nucleophilic reaction. In certain embodiments, Q comprises a click probe, a thiol, a thiol -reactive moiety, an amine or an amine-reactive moiety. In certain embodiments, Q is a click probe, a thiol-reactive moiety or an amine-reactive moiety. In certain embodiments, Q is a click probe. In certain embodiments, the click probe is reactive in a cycloaddition (click reaction). In certain embodiments, the click probe is selected from an azide, a tetrazine, a triazine, a nitrone, a nitrile oxide, a nitrile imine, a diazo compound, an or / Ao-quinone, a dioxothiophene, a sydnone, an alkene moiety and an alkyne moiety. In certain embodiments, the click probe comprises or is an alkene moiety or an alkyne moiety. In certain embodiments, the alkene is a (hetero)cycloalkene and / or the alkyne is a terminal alkyne or a (hetero)cycloalkyne. Typical thiol -reactive moieties are selected from maleimide moiety, a haloacetamide moiety, an allenamide moiety, a phosphonamidite moiety, a cyanoethynyl moiety, a vinylsulfone, a vinylpyridine moiety or a methylsulfonylphenyloxadiazole moiety. In certain embodiments, the thiol -reactive moiety comprises or is a maleimide moiety. Typical amine-reactive moieties are selected from N-hydroxysuccinimidyl esters and other activated esters, / >-nitrophenyl carbonates and other activated carbonates, isocyanates, isothiocyanates, haloacetamides and benzyl halides. In certain embodiments, Q is selected from an alkene moiety, an alkyne moiety, a thiol-reactive moiety and an amine-reactive moiety. In certain embodiments, Q is selected from an alkene moiety and an alkyne moiety. In certain embodiments, Q is an alkyne moiety. In certain embodiments, the alkene is a (hetero)cycloalkene and the alkyne is a terminal alkyne or a (hetero)cycloalkyne. In certain embodiments, Q is a cyclic (hetero)alkyne moiety. Each of these moieties are further defined here below.

[0268] In certain embodiments, Q comprises a cyclic (hetero)alkyne moiety. The alkynyl group may also be referred to as a (hetero)cycloalkynyl group, / .e., a heterocycloalkynyl group ora cycloalkynyl group, wherein the (hetero)cycloalkynyl group is optionally substituted. In certain embodiments, the (hetero)cycloalkynyl group is a (hetero)cycloheptynyl group, a (hetero)cyclooctynyl group, a (hetero)cyclononynyl group or a (hetero)cyclodecynyl group. In certain embodiments, the (hetero)cycloalkynes are optionally substituted. In certain embodiments, the (hetero)cycloalkynyl group is an optionally substituted (hetero)cycloheptynyl group or an optionally substituted (hetero)cyclooctynyl group. In certain embodiments, the (hetero)cycloalkynyl group is a (hetero)cyclooctynyl group, wherein the (hetero)cyclooctynyl group is optionally substituted.

[0269] In certain embodiments, Q comprises a (hetero)cycloalkynyl or (hetero)cycloalkenyl group and is according to structure (QI):wherein:the bond depicted as - is a double bond or a triple bond;R15is independently selected from the group consisting of hydrogen, halogen, -OR16, -NO2, -CN, -S(O)2R16, -S(O)3(-), CI - C24 alkyl groups, Ce - C24 (hetero)aryl groups, C7 - C24 alkyl(hetero)aryl groups and C7 - C24 (hetero)aryl alkyl groups and wherein the alkyl groups, (hetero)aryl groups, alkyl(hetero)aryl groups and (hetero)arylalkyl groups are optionally substituted, wherein two substituents R15may be linked together to form an optionally substituted annulated cycloalkyl or an optionally substituted annulated (hetero)arene substituent, and wherein R16is independently selected from the group consisting of hydrogen, halogen, Ci - C24 alkyl groups, Ce - C24 (hetero)aryl groups, C7 - C24 alkyl(hetero)aryl groups and C7 - C24 (hetero)aryl alkyl groups;Y2is C(R31)2, O, S, S(+)R31, S(O)R31, S(O)=NR31or NR31, wherein S(+)is a cationic sulphur atom counterbalanced by B(_), wherein B( )is an anion, and wherein each R31individually is R15or a connection with D, connected via L;u is 0, 1, 2, 3, 4 or 5;u’ is 0, 1, 2, 3, 4 or 5, wherein u + u’ = 0, 1, 2, 3, 4, 5, 6, 7 or 8;v = an integer in the range 0 - 16.

[0270] Typically, v = (u + u’) x 2 (when the connection to L, depicted by the wavy bond, is via Y2) or [(u + u’) x 2] - 1 (when the connection to L, depicted by the wavy bond, is via one of the carbon atoms).

[0271] In certain embodiments of structure (QI), reactive group Q comprises a (hetero)cycloalkynyl group and is according to structure (QI a):wherein,R15and Y2are as defined aboveu is 0, 1, 2, 3, 4 or 5;u’ is 0, 1, 2, 3, 4 or 5, wherein u + u’ = 4, 5, 6, 7 or 8;v = an integer in the range 8 - 16.

[0272] In certain embodiments, u + u’ = 4, 5 or 6. In certain embodiments, u + u’ = 5. In certain embodiments, v = 8, 9 or 10. In certain embodiments, v = 9 or 10. In certain embodiments, v = 10.

[0273] In certain embodiments, Q is a (hetero)cycloalkynyl group selected from the group consisting of (Q2) - (Q20) and (Q20a) depicted here below.wherein the connection to L, depicted with the wavy bond, may be to any available carbon or nitrogen atom of Q. The nitrogen atom of (Q 10), (Q13), (Q14) and (Q 15) may bear the connection to L, or may comprise a hydrogen atom or be optionally functionalized. B(’ is an anion, which is in certain embodiments selected fromf’OTf. CT Br*^ or f In certain embodiments, B(- is(Tf. In the conjugation reaction, B(' does not need to be a pharmaceutically acceptable anion, since Bf’ will exchange with the anions present in the reaction mixture anyway. In certain embodiments, in case (Q 19) is used for Q, the negatively charged counter-ion is pharmaceutically acceptable upon isolation of the conjugate according to the present disclosure, such that the conjugate is readily useable as medicament.

[0274] In certain embodiments, Q is a (hetero)cycloalkynyl group selected from the group consisting of (Q21 ) - (Q38) and (Q38a) depicted here below.

[0275] In structure (Q38), B(is an anion, which is, e.g., selected from(Tf, Cl(Br( )or I(In certain embodiments, B(is(^Tf

[0276] In certain embodiments, Q comprises a (hetero) cyclooctyne moiety or a (hetero)cycloheptyne moiety, e.g., according to structure (Q8), (Q26), (Q27), (Q28), (Q37) or (Q38a) or according to structure (Q8), (Q26), (Q27), (Q28) or (Q37), which are optionally substituted. Each of these options for Q are further defined here below.

[0277] In certain embodiments, Q comprises a heterocycloheptyne moiety according to structure (Q37), also referred to as a TMTHSI, which is optionally substituted. In certain embodiments, the heterocycloheptyne moiety according to structure (Q37) is not substituted.

[0278] In certain embodiments, Q comprises a cyclooctyne moiety according to structure (Q8) or to structure (Q29), also referred to as abicyclo[6.1.0]non-4-yn-9-yl] group (BCN group), which is optionally substituted. In certain embodiments, the cyclooctyne moiety according to structure (Q8) or (Q29) is not substituted. In certain embodiments, Q is a (hetero)cyclooctyne moiety according to structure (Q39) as shown below, wherein V is (CH2)I and 1 is an integer in the range of 0 to 10, e.g., in the range of 0 to 6. In certain embodiments, 1 is 0, 1, 2, 3 or 4. In certain embodiments, 1 is 0, 1 or 2. In certain embodiments, 1 is 0 or 1. In certain embodiments of group (Q39), 1 is 1. In certain embodiments, Q is according to structure (Q42), defined further below.

[0279] In certain embodiments, Q comprises a (hetero)cyclooctyne moiety according to structure (Q26), (Q27) or (Q28), also referred to as a DIBO, DIBAC, DBCO or ADIBO group, which are optionally substituted. In certain embodiments, Q is a (hetero)cyclooctyne moiety according to structure (Q40) or (Q41) as shown below, wherein Y1is O or NR11, wherein R11is independently selected from the group consisting of hydrogen, a linear or branched Ci - Cn alkyl group or a C4 - C12 (hetero)aryl group. The aromatic rings in (Q40) are optionally O-sulfonylated at one or more positions, whereas the rings of (Q41) may be halogenated at one or more positions. In certain embodiments, the (hetero)cyclooctyne moiety according to structure (Q40) or (Q41) is not further substituted. In certain embodiments, Q is according to structure (Q43), defined further below.

[0280] In certain embodiments, Q comprises a heterocycloheptynyl group and is according to structure (Q37).

[0281] In certain embodiments, Q comprises a cyclooctynyl group and is according to structure (Q42):wherein:R15is independently selected from the group consisting of hydrogen, halogen, -OR16, -NO2, -CN, -S(O)2R16, -S(O)3('), CI - C24 alkyl groups, C5 - C24 (hetero)aryl groups, C7 - C24 alkyl(hetero)aryl groups and C7 - C24 (hetero)aryl alkyl groups and wherein the alkyl groups, (hetero)aryl groups, alkyl(hetero)aryl groups and (hetero)arylalkyl groups are optionally substituted, wherein two substituents R15may be linked together to form an optionally substituted annulated cycloalkyl or an optionally substituted annulated (hetero)arenesubstituent, and wherein R16is independently selected from the group consisting of hydrogen, halogen, Ci - C24 alkyl groups, Ce - C24 (hetero)aryl groups, C7 - C24 alkyl(hetero)aryl groups and C7 - C24 (hetero)aryl alkyl groups;R18is independently selected from the group consisting of hydrogen, halogen, Ci - C24 alkyl groups, Ce - C24 (hetero)aryl groups, C7 - C24 alkyl(hetero)aryl groups and C7 - C24 (hetero)arylalkyl groups;R19is selected from the group consisting of hydrogen, halogen, Ci - C24 alkyl groups, Ce - C24 (hetero)aryl groups, C7 - C24 alkyl(hetero)aryl groups and C7 - C24 (hetero)arylalkyl groups, the alkyl groups optionally being interrupted by one of more hetero-atoms selected from the group consisting of O, N and S, wherein the alkyl groups, (hetero)aryl groups, alkyl(hetero)aryl groups and (hetero)arylalkyl groups are independently optionally substituted, or R19is a second occurrence of Q or D connected via a spacer moiety; and1 is an integer in the range 0 to 10.

[0282] In certain embodiments of the reactive group according to structure (Q42), R15is independently selected from the group consisting of hydrogen, halogen, -OR16, Ci - Ce alkyl groups, C5 - Ce (hetero)aryl groups, wherein R16is hydrogen or Ci - Ce alkyl. In certain embodiments, R15is independently selected from the group consisting of hydrogen and Ci - Ce alkyl. In certain embodiments, all R15are H. In certain embodiments of the reactive group according to structure (Q42), R18is independently selected from the group consisting of hydrogen, Ci - Ce alkyl groups. In certain embodiments, both R18are H. In certain embodiments of the reactive group according to structure (Q42), R19is H. In certain embodiments of the reactive group according to structure (Q42), I is 0 or 1. In certain embodiments, 1 is 1.

[0283] In certain embodiments, Q comprises a (hetero)cyclooctynyl group and is according to structure (Q43):wherein:R15is independently selected from the group consisting of hydrogen, halogen, -OR16, -NO2, -CN, -S(O)2R16, -S(O)3(’), CI - C24 alkyl groups, C5 - C24 (hetero)aryl groups, C7 - C24alkyl(hetero)aryl groups and C7 - C24 (hetero)aryl alkyl groups and wherein the alkyl groups, (hetero)aryl groups, alkyl(hetero)aryl groups and (hetero)arylalkyl groups are optionally substituted, wherein two substituents R15may be linked together to form an optionally substituted annulated cycloalkyl or an optionally substituted annulated (hetero)arene substituent, and wherein R16is independently selected from the group consisting of hydrogen, halogen, Ci - C24 alkyl groups, Ce - C24 (hetero)aryl groups, C7 - C24 alkyl(hetero)aryl groups and C7 - C24 (het ero)aryl alkyl groups;Y is N or CR15.

[0284] In certain embodiments of the reactive group according to structure (Q43), R15is independently selected from the group consisting of hydrogen, halogen, -OR16, -S(O)3(-), Ci - Ce alkyl groups, C5 - Ce (hetero)aryl groups, wherein R16is hydrogen or Ci - Ce alkyl. In certain embodiments, R17is independently selected from the group consisting of hydrogen and -S(O)3(-). In certain embodiments of the reactive group according to structure (Q43), Y is N or CH. In certain embodiments, Y = N.

[0285] In certain embodiments, Q comprises a heterocycloheptynyl group and is according to structure (Q37) or (Q38a). In certain embodiments, Q comprises a heterocycloheptynyl group and is according to structure (Q37).(Q37) (Q38a)

[0286] In certain embodiments, Q comprises a cyclic alkene moiety. The alkenyl group Q may also be referred to as a (hetero)cycloalkenyl group, i.e., a heterocycloalkenyl group or a cycloalkenyl group, c.., a cycloalkenyl group, wherein the (hetero)cycloalkenyl group is optionally substituted. In certain embodiments, the (hetero)cycloalkenyl group is a (hetero)cyclopropenyl group, a (hetero)cyclobutenyl group, a norbomene group, a norbomadiene group, a / ra / .s-(hetero)cycloheptenyl group, a / ra / 7.s-(hetero)cyclooctenyl group, a trans-(hetero)cyclononenyl group or a Zra / / .s-(hetero)cyclodecenyl group, which may all optionally be substituted. In certain embodiments, the (hetero)cycloalkenyl group is a (hetero)cyclopropenyl group, a / / zw7.s-(hetero)cycloheptenyl group, or a / / z7 / 7.s-(hetero)cyclooctenyl group, wherein the (hetero)cyclopropenyl group, the / / •c / / 7.s-(hetero)cycloheptenyl group or the trans-(hetero)cyclooctenyl group is optionally substituted. In certain embodiments, Q comprises a cyclopropenyl moiety according to structure (Q44), a hetereocyclobutene moiety according to structure (Q45), a norbornene or norbomadiene group according to structure (Q46), a trans- (hetero)cycloheptenyl moiety according to structure (Q47) or a / raw.s,-(hetero)cyclooctenyl moiety according to structure (Q48). Herein, Y3is selected from C(R23)2, NR23or O, wherein each R23is individually hydrogen, Ci - Ce alkyl or is connected to L, optionally via a spacer, and the bond labelled - is a single or double bond. In certain embodiments, the cyclopropenyl group is according to structure (Q49). In certain embodiments, the / / / / 7.s-(heteio)cycloheptene group is according to structure (Q50) or (Q51). In certain embodiments, the / ra / z.s-(hetero)cyclooctene group is according to structure (Q52), (Q53 ), (Q54), (Q55) or (Q56).wherein the R group(s) on Si in (Q50) and (Q51) are typically alkyl or aryl, e.g., Ci-Ce alkyl.

[0287] In certain embodiments, Q is a thiol -reactive probe. In such embodiments, Q is a reactive group compatible with cysteine conjugation. Such probes are known in the art and may be selected from the group consisting of a maleimide moiety, a haloacetamide moiety, an allenamide moiety, a phosphonamidite moiety, a cyanoethynyl moiety, a vinylsulfone, a vinylpyridine moiety or a methylsulfonylphenyloxadiazole moiety. In certain embodiments, Q comprises or is a maleimide moiety. Reagents may be monoalkylation type or may be a crosslinker for reaction with two cysteine side-chains.

[0288] In certain embodiments, probe Q is selected from the group consisting of (Q57) - (Q71) depicted here below.wherein:X6is H, halogen, PhS, MeS, e.g., a halogen, such as Cl, Br, I;X7is halogen, PhS, MeS, e.g., a halogen, such as Cl, Br, I;R24is H or Ci-12 alkyl, e.g., H or Ci-6 alkyl;R25is H, C 1-12 alkyl, Ci-12 aryl, C1-12 alkaryl or C1-12 aralkyl, e.g., H or jczra-methylphenyl; wherein the aromatic ring of (Q61) and (Q63) may optionally be a heteroaromatic ring, such as a phenyl or pyridine ring.

[0289] In certain embodiments of thiol -reactive probe (Q57), the probe Q is selected from the group consisting of (Q72) - (Q74) depicted here below.(Q72) (Q73) (Q74)wherein:R27is Ci-12 alkyl, C1.12 aryl, C1-12 alkaryl or C1-12 aralkyl;t is an integer in the range of 0 - 15, e.g., 1 - 10.

[0290] In certain embodiments, Q is an amine-reactive probe. In this embodiment, Q is a reactive group compatible with lysine conjugation. Such probes are known in the art and may be selected from the group consisting of A -hydroxy sue ni mi dyl groups, isocyanate groups, isothiocyanate groups and benzoyl halide groups. In certain embodiments, Q comprises or is an N-hydroxysuccinimidyl esters or a p-nitrophenyl carbonate moiety.

[0291] In certain embodiments, probe Q is selected from the group consisting of (Q75) - (Q79) depicted here below.(Q75) (Q76) (Q77) (Q78) (Q79)wherein X2is a halogen, e.g., F.

[0292] In certain embodiments, Q is selected from the group consisting of (QI) - (Q79).

[0293] In certain embodiments, an ADC provided herein is synthesized, e.g., as described below, using a compound of general structure (2), wherein L-D (the linker-payload moiety) is as in ADC embodiment (Xllg), wherein payload D is MMAE and Q is a click probe. In certain embodiments, Q is a (hetero)cyclooctyne moiety according to structure (Q39), wherein V = CH2. In certain embodiments, Q is according to structure (Q42), wherein all R17, all R18, and R19are each hydrogen and 1 is 1. In certain embodiments, an ADC provided herein is synthesized, e.g., as described below, using a compound of general structure (2), wherein the compound of general structure (2) is as shown in FIG.3A.

[0294] In certain embodiments, an ADC provided herein is synthesized, e.g., as described below, using a compound of general structure (2), wherein L-D (the linker-payload moiety) is as in ADC embodiment (XHIe), wherein payload D is exatecan and Q is a click probe. In certain embodiments, Q is a (hetero)cyclooctyne moiety according to structure (Q39), wherein V = CH2. In certain embodiments, Q is according to structure (Q42), wherein all R15, all R18, and R19are each hydrogen and 1 is 1. In certain embodiments, an ADC provided herein is synthesized, e.g., as described below, using a compound of general structure (2), wherein the compound of general structure (2) is as shown in FIG.3B.Antibody according to general structure (3)

[0295] In certain embodiments, the antibody of the ADCs provided herein has general structure (3):AB-[(L6)b-{F}x]y(3)wherein:AB is an antibody that specifically binds to LY6K (e.g., human LY6K);b is 0 or 1;L6is -GlcNAc(Fuc)w-(G)j-S-(L7)W’-, wherein G is a monosaccharide, j is an integer in the range of 0 - 10, S is a sugar or a sugar derivative, GlcNAc is N-acetylglucosamine and Fuc is fucose, w is 0 or 1, w’ is 0, 1 or 2 and L7is -N(H)C(O)CH2-, -N(H)C(O)CF2- or -CH2-;F is a reactive moiety;x is 1 or 2; andy is 1, 2, 3 or 4.

[0296] The antibody of general structure (3) may also be referred to as a “(modified) antibody”, for being an antibody comprising reactive groups F, wherein the reactive groups F are naturally present, or the antibody is modified to incorporate the reactive groups F. The (modified) antibody according to general structure (3) can be prepared by the skilled person according to the Examples herein and / or using standard organic and / or enzymatic synthesis techniques. Antibody AB, linker L6, b, x and y are defined above in the context of the conjugate according to structure (1).Reactive moiety F

[0297] F is reactive towards Q in the conjugation reaction defined below, e.g., wherein the conjugation reaction is a cycloaddition or a nucleophilic reaction. As the skilled person will understand, the options for F are the same as those for Q, provided that F and Q are reactive towards each other. In certain embodiments, F comprises a click probe, a thiol, a thiol-reactive moiety, an amine or an amine-reactive moiety. In certain embodiments, F is a click probe, a thiol or an amine. In certain embodiments, F is a click probe. In certain embodiments, the click probe is reactive in a cycloaddition (click reaction). In certain embodiments, the click probe is selected from an azide, a tetrazine, a triazine, a nitrone, a nitrile oxide, a nitrile imine, a diazo compound,an or / Ao-quinone, a dioxothiophene, a sydnone, an alkene moiety and an alkyne moiety. In certain embodiments, the click probe comprises or is an azide, a tetrazine, a triazine, a nitrone, a nitrile oxide, a nitrile imine, a diazo compound, an or / Ao-quinone, a dioxothiophene or a sydnone. In certain embodiments, the click probe is an azide. Typical thiol -reactive moieties are selected from maleimide moiety, a haloacetamide moiety, an allenamide moiety, a phosphonamidite moiety, a cyanoethynyl moiety, an ort / zo-quinone moiety, a vinylsulfone, a vinylpyridine moiety or a methylsulfonylphenyloxadiazole moiety. In certain embodiments, the thiol -reactive moiety comprises or is a maleimide moiety. Typical amine-reactive moieties are selected from N-hydroxysuccinimidyl esters, isocyanates, isothiocyanates and benzyl halides. In certain embodiments, F is a click probe or a thiol. In certain embodiments, F is an azide or a thiol. In certain embodiments, F is an azide.

[0298] More than one reactive group F may be present in the antibody. The reactive group F in the antibody may be naturally present or may be placed in the antibody by a specific technique, for example a (bio)chemical or a genetic technique. In certain embodiments, the reactive group that is placed in the antibody is prepared by chemical synthesis, e.g., an azide or a terminal alkyne. Methods of preparing modified antibodies are known in the art, e.g., from WO 2014 / 065661, WO 2016 / 170186 and WO 2016 / 053107, which are incorporated herein by reference. From the same documents, the conjugation reaction between the modified antibody and a linker-drug construct is known to the skilled person.

[0299] In certain embodiments, F is a click probe reactive towards a (hetero)cycloalkene and / or a (hetero)cycloalkyne. In certain embodiments, F is selected from the group consisting of azide, tetrazine, triazine, nitrone, nitrile oxide, nitrile imine, diazo compound, or / Ao-quinone, di oxothiophene and sydnone. In certain embodiments, the reactive moiety F is selected from structures (Fl) - (F 10) depicted here below.wherein the wavy bond represents the connection to the payload. For (F3), (F4), (F8) and (F9), the payload can be connected to any one of the wavy bonds. The other wavy bond may then be connected to an R group selected from hydrogen, Ci - C24 alkyl groups, C2 - C24 acyl groups, C3 - C24 cycloalkyl groups, C2 - C24 (hetero)aryl groups, C3 - C24 alkyl(hetero)aryl groups, C3 - C24 (hetero)arylalkyl groups and Ci - C24 sulfonyl groups, each of which (except hydrogen) may optionally be substituted and optionally interrupted by one or more heteroatoms selected from O, S and NR32wherein R32is independently selected from the group consisting of hydrogen and Ci - C4 alkyl groups. The skilled person understands which R groups may be applied for each of the groups F. For example, the R group connected to the nitrogen atom of (F3) may be selected from alkyl and aryl, and the R group connected to the carbon atom of (F3) may be selected from hydrogen, alkyl, aryl, acyl and sulfonyl. In certain embodiments, the reactive moiety F is selected from azides or tetrazines. In certain embodiments, the reactive moiety F is an azide.

[0300] In certain embodiments, F is a thiol or precursor thereof. Thiol or precursor thereof F is used in the conjugation reaction to connect the linker-drug construct to the (modified) antibody. F is reactive towards thiol-reactive probe Q in a thiol ligation. The thiol, e.g., the thiol of the side chain of a cysteine amino acid, may be naturally present within the antibody AB, in which case linker L6is not present (b = 0), although it may also be synthetically introduced, optionally via a linker L6. Thiol precursors in the context of bioconjugation are known in the art, and include disulfides, which may be naturally occurring disulfide bridges present in the antibody or synthetically introduced disulfides, which are reduced as known in the art. In certain embodiments, F is a thiol group of a cysteine side chain.

[0301] In certain embodiments, F is an amine or precursor thereof, e.g., an amine. Amine or precursor thereof F is used in the conjugation reaction to connect the linker-drug construct to the (modified) antibody. F is reactive towards amine-reactive probe Q in nucleophilic substitution. The amine is typically a primary amine, e.g., the amine of the side chain of a lysine amino acid, which are naturally present within the antibody AB, in which case linker L6is not present (b = 0), although it may also be synthetically introduced, optionally via a linker L6. In certain embodiments, F is a primary amine group of a lysine side chain.Process for synthesising the ADC according to general structure fl)

[0302] In certain embodiments, the ADCs of the present disclosure can be prepared according to a process comprising the step of reacting Q of the compound according to the invention with areactive group F of an antibody. The compound according to general structure (2), and embodiments thereof, are described in more detail above. The present process occurs under conditions such that Q is reacted with F to covalently link the antibody AB (3) to the payload D. In the process according to the invention, Q reacts with F, forming a covalent connection between the antibody and the compound according to the invention. Complementary reactive groups Q and reactive groups F are known to the skilled person and are described in more detail below.

[0303] Any conjugation technique known in the art can be employed to prepare the multifunctional antibody constructs according to the invention. Suitable conjugation techniques include thiol ligation, lysine ligation, cycloadditions (e.g., copper-catalysed click reaction, strain-promoted azide-alkyne cycloaddition, strain-promoted quinone-alkyne cycloaddition). In certain embodiments, conjugation techniques useful in the context of the present disclosure include nucleophilic reactions and cycloadditions, e.g., wherein the cycloaddition is a [4+2] cycloaddition or a [3+2] cycloaddition and the nucleophilic reaction is a Michael addition or a nucleophilic substitution. Suitable conjugation techniques are disclosed in, e.g, G. T. Hermanson, “Bioconjugate Techniques”, Elsevier, 3rd Ed. 2013 (ISBN:978-0-12-382239-0), WO 2014 / 065661, van Geel et al., Bioconj. Chern. 2015, 26, 2233-2242, PCT / EP2021 / 050594, PCT / EP2021 / 050598 and NL 2026947.

[0304] In certain embodiments, conjugation is accomplished via a nucleophilic reaction, such as a nucleophilic substitution or a Michael reaction. In certain embodiments, the nucleophilic reaction is the acylation of a primary amino group with an activated ester. In certain embodiments, the Michael reaction is the maleimide-thiol reaction, which is widely employed in bioconjugation.

[0305] In certain embodiments, conjugation is accomplished via a cycloaddition. In certain embodiments, the cycloaddition is a (4+2)-cycloaddition (e.g., a Diels- Alder reaction) or a (3+2)-cycloaddition (e.g., a 1,3-dipolar cycloaddition). In certain embodiments, the conjugation reaction is the Diels-Alder reaction or the 1,3-dipolar cycloaddition. In certain embodiments, the Diels-Alder reaction is the inverse-electron demand Diels-Alder cycloaddition. In certain embodiments, the 1,3-dipolar cycloaddition is used, e.g., the alkyne-azide cycloaddition. In certain embodiments, Q is or comprises an alkyne group and F is an azido group. Cycloadditions, such as Diels-Alder reactions and 1,3-dipolar cycloadditions are known in the art, and the skilled person knows how to perform them.

[0306] In certain embodiments, the process for preparing an ADC provided herein comprises a click reaction, e.g., a 1,3 -dipolar cycloaddition, or an alkyne / azide cycloaddition. In certain embodiments, Q is or comprises an alkyne group and F is an azido group. Click reactions, such as 1,3 -dipolar cycloadditions, are known in the art, and the skilled person knows how to perform them.

[0307] In certain embodiments, the process for preparing an ADC provided herein comprises reacting the modified antibody of structure (3) with a compound according to structure (2), to obtain the ADC of structure (1).

[0308] In certain embodiments, the process for preparing an ADC provided herein comprises:(i) contacting an antibody comprising y core / V-acetylglucosamine (GlcNAc) moieties, wherein y = 1, 2, 3 or 4, with a compound of the formula S(F)X-P in the presence of a catalyst, wherein S(F)Xis a sugar derivative comprising x reactive groups F capable of reacting with a reactive group Q, x is 1 or 2 and P is a nucleoside mono- or diphosphate, and wherein the catalyst is capable of transferring the S(F)Xmoiety to the core-GlcNAc moiety, to obtain a modified antibody according to Formula (26):AB-[GlcNAc(Fuc)w-S {F }x]y(26)whereinAB is an antibody that specifically binds to LY6K (e.g., human LY6K);Fuc is fucose;w is 0 or 1; and(ii) reacting the modified antibody with a compound according to structure (2):Q-L-D(2)wherein:Q is a reactive moiety;L is a linker that links Z to D;D is selected from the group consisting of taxanes, anthracyclines, camptothecins, epothilones, mytomycins, combretastatins, vinca alkaloids, tubulysins, enediynes (e.g., calicheamicins), amanitins, duocarmycins, maytansinoids, amatoxins, dolastatins, auristatins, eribulins, BCL-XL inhibitors, hemiasterlins, KSP inhibitors, radioisotopes, TLR agonists,indolinobenzodi azepine dimers or pyrrolobenzodiazepine dimers (PBDs), and analogues or prodrugs thereof;to obtain the ADC according to structure (1).Step (i)

[0309] In step (i), an antibody comprising 1, 2, 3 or 4 core N-acetylglucosamine moi eties is contacted with a compound of the formula S(F)X-P in the presence of a catalyst, wherein S(F)Xis a sugar derivative comprising x reactive groups F capable of reacting with a reactive group Q, x is 1 or 2 and P is a nucleoside mono- or diphosphate, and wherein the catalyst is capable of transferring the S(F)Xmoiety to the core-GlcNAc moiety. Herein, the antibody is typically an antibody that has been trimmed to a core-GlcNAc residue as described further below. Step (i) affords a modified antibody according to Formula (26).

[0310] The starting material, i.e., the antibody comprising a core-GlcNAc substituent, is known in the art and can be prepared by methods known by the skilled person. In one embodiment, the process described herein further comprises the deglycosylation of an antibody glycan having a core N-acetylglucosamine, in the presence of an endoglycosidase, in order to obtain an antibody comprising a core N-acetylglucosamine substituent, wherein said core N-acetylglucosamine and said core N-acetylglucosamine substituent are optionally fucosylated. Depending on the nature of the glycan, a suitable endoglycosidase may be selected. In certain embodiments, the endoglycosidase is selected from the group consisting of EndoS, EndoA, EndoE, EfEndol8A, EndoF, EndoM, EndoD, EndoH, EndoT and EndoSH and / or a combination thereof, the selection of which depends on the nature of the glycan. EndoSH is described in PCT / EP2017 / 052792, see Examples 1 - 3, and SEQ. ID No: 1, which is incorporated by reference herein.

[0311] Structural features S and x are defined above for the ADC provided herein, which equally applies to the present aspect. Compounds of the formula S(F)X-P, wherein a nucleoside monophosphate or a nucleoside diphosphate P is linked to a sugar derivative S(F)X, are known in the art. For example, Wang et al., Chem. Eur. J. 2010, 16, 13343-13345, Piller et al., ACS Chem. Biol. 2012, 7, 753, Piller et al., Bioorg. Med. Chem. Lett. 2005, 15, 5459-5462 and WO 2009 / 102820, all incorporated by reference herein, disclose a number of compounds S(F)X-P and their syntheses. In certain embodiments, nucleoside mono- or diphosphate P in S(F)X-P is selected from the group consisting of uridine diphosphate (UDP), guanosine diphosphate (GDP), thymidine diphosphate (TDP), cytidine diphosphate (CDP) and cytidine monophosphate (CMP). In certainembodiments, P is selected from the group consisting of uridine diphosphate (UDP), guanosine diphosphate (GDP) and cytidine diphosphate (CDP). In certain embodiments, P = UDP. In certain embodiments, S(F)X-P is selected from the group consisting of GalNAz-UDP, F2-GalNAz-UDP (A-(azidodifluoro)acetyl-galactosamine), 6-AzGal-UDP, 6-AzGalNAc-UDP (6-azido-6-deoxy-N-acetylgalactosamine-UDP), 4-AzGalNAz-UDP, 6-AzGalNAz-UDP, GlcNAz-UDP, 6-AzGlc-UDP, 6-AzGlcNAz-UDP and 2-(but-3-yonic acid amido)-2-deoxy-galactose-UDP. In certain embodiments, S(F)X-P is GalNAz-UDP or 6-AzGalNAc-UDP.

[0312] Suitable catalyst that are capable of transferring the S(F)Xmoiety to the core-GlcNAc moiety are known in the art. A suitable catalyst is a catalyst wherefore the specific sugar derivative nucleotide S(F)X-P in that specific process is a substrate. More specifically, the catalyst catalyses the formation of a P(l,4)-glycosidic bond. In certain embodiments, the catalyst is selected from the group of galactosyltransferases and N-acetylgalactosaminyltransferases. In certain embodiments, the catalyst is selected from the group of P(l,4)-N-acetylgalactosaminyltransferases (GalNAcT) and P(l,4)-galactosyltransf erases (GalT). In certain embodiments, the catalyst is selected from the group of P(l,4)-N-acetylgalactosaminyltransferases having a mutant catalytic domain. Suitable catalysts and mutants thereof are disclosed in WO 2014 / 065661, WO 2016 / 022027 and WO 2016 / 170186, all incorporated herein by reference. In certain embodiments, the catalyst is a wild-type galactosyltransferase or N-acetylgalactosaminyltransferase. In certain embodiments, the catalyst is a wild-type N-acetylgalactosaminyltransferase. In certain embodiments, the catalyst is a mutant galactosyltransferase orN-acetylgalactosaminyltransferases. In certain embodiments, the catalyst is a mutant N-acetylgalactosaminyltransferase. In certain embodiments, the catalyst is a mutant enzyme described in WO 2016 / 022027 and WO 2016 / 170186. These galactosyltransferase (mutant) enzyme catalysts are able to recognize internal sugars and sugar derivatives as an acceptor. Thus, sugar derivative S(F)Xis linked to the core-GlcNAc substituent in step (i), irrespective of whether said GlcNAc is fucosylated or not.

[0313] In certain embodiments, Step (i) is performed in a suitable buffer solution, such as for example phosphate, buffered saline (e.g., phosphate-buffered saline, tris-buffered saline), citrate, HEPES, tris and glycine. Suitable buffers are known in the art. In certain embodiments, the buffer solution is phosphate-buffered saline (PBS) or tris buffer. In certain embodiments, Step (i) is performed at a temperature in the range of about 4 to about 50 °C, e.g., in the range of about 10 to about 45 °C, in the range of about 20 to about 40 °C, or in the range of about 30 to about 37 °C.In certain embodiments, Step (i) is performed at a pH in the range of about 5 to about 9, e.g., in the range of about 5.5 to about 8.5, in the range of about 6 to about 8, or in the range of about 7 to about 8.Step (ii)

[0314] In step (ii), the modified antibody is reacted with a compound according to general structure (2), comprising a reactive group Q capable of reacting with reactive group F and a payload D, to obtain the ADC according to structure (1), comprising connecting group Z resulting from the reaction between Q and F. Such reaction occurs under condition such that reactive group Q is reacted with the reactive group F of the biomolecule to covalently link the antibody to the compound according to general structure (2). Step (ii) may also be referred to as the conjugation reaction.

[0315] In certain embodiments, in Step (ii) an azide on an azide-modified antibody reacts with an alkynyl group, e.g., a terminal alkynyl group, or a (hetero)cycloalkynyl group of the compound according to general structure (2), via a cycloaddition reaction. This cycloaddition reaction of a molecule comprising an azide with a molecule comprising a terminal alkynyl group or a (hetero)cycloalkynyl group is one of the reactions that is known in the art as “click chemistry”. In the case of a linker-conjugate comprising a terminal alkynyl group, said cycloaddition reaction needs to be performed in the presence of a suitable catalyst, e.g., a Cu(I) catalyst. In certain embodiments, the linker-conjugate comprises a (hetero)cycloalkynyl group, e.g, a strained (hetero)cycloalkynyl group. When the (hetero)cycloalkynyl is a strained (hetero)cycloalkynyl group, the presence of a catalyst is not required, and said reaction may even occur spontaneously by a reaction called strain -promoted azide-alkyne cycloaddition (SPAAC). This is one of the reactions known in the art as “metal-free click chemistry”.Mode of conjugation

[0316] In the context of the present disclosure, the “mode of conjugation” refers to the process that is used to conjugate a payload D to an antibody AB, as well as to the structural features of the resulting ADC, in particular of the linker that connects the payload to the antibody, that are a direct consequence of the process of conjugation. Thus, in one embodiment, the mode of conjugation refers to a process for conjugation of a payload to an antibody. In an alternative embodiment, the mode of conjugation refers to structural features of the linker and / or to the attachment point of thelinker to the antibody that are a direct consequence of the process for conjugation a payload to an antibody.

[0317] In a further aspect, the present disclosure concerns the use of a mode of conjugation for increasing the therapeutic index of an ADC in the methods of treatment provided herein, wherein the mode of conjugation is being used to connect antibody AB with payload D via a linker L. In certain embodiments, the mode of conjugation comprises:(i) contacting an antibody comprising y core A-acetylglucosamine (GlcNAc) moi eties, wherein y = 1, 2, 3 or 4, with a compound of the formula S(F)X-P in the presence of a catalyst, wherein S(F)Xis a sugar derivative comprising x reactive groups F capable of reacting with a reactive group Q, x is 1 or 2 and P is a nucleoside mono- or diphosphate, and wherein the catalyst is capable of transferring the S(F)Xmoiety to the core-GlcNAc moiety, to obtain a modified antibody according to Formula (26):AB-[GlcNAc(Fuc)w-S {F }x]y(26)whereinAB is an antibody that specifically binds to LY6K (e.., human LY6K);Fuc is fucose;w is 0 or 1; and(ii) reacting the modified antibody with a compound according to structure (2):Q-L-D(2)wherein:Q is a reactive moiety;L is a linker that links Z to D;D is selected from the group consisting of taxanes, anthracyclines, camptothecins, epothilones, mytomycins, combretastatins, vinca alkaloids, tubulysins, enediynes (e.g., calicheamicins), amanitins, duocarmycins, maytansinoids, amatoxins, dolastatins, auristatins, eribulins, BCL-XL inhibitors, hemiasterlins, KSP inhibitors, radioisotopes, TLR agonists, indolinobenzodiazepine dimers or pyrrolobenzodiazepine dimers (PBDs), and analogues or prodrugs thereof;to obtain the antibody-conjugate according to structure (1)

[0318] In certain embodiments, increasing the therapeutic index of an ADC is selected from: (a) increasing the therapeutic efficacy of the ADC; and / or(b) increasing the tolerability of the ADC.Pharmaceutical Compositions

[0319] Provided herein are compositions comprising an ADC disclosed herein having the desired degree of purity in a physiologically acceptable carrier, excipient, or stabilizer (see, e.g., Remington’s Pharmaceutical Sciences (1990) Mack Publishing Co., Easton, PA). Acceptable carriers, excipients, or stabilizers are nontoxic to recipients at the dosages and concentrations employed, and include buffers such as phosphate, citrate, and other organic acids; antioxidants including ascorbic acid and methionine; preservatives (such as octadecyldimethylbenzyl ammonium chloride; hexamethonium chloride; benzalkonium chloride, benzethonium chloride; phenol, butyl or benzyl alcohol; alkyl parabens such as methyl or propyl paraben; catechol; resorcinol; cyclohexanol; 3-pentanol; and m-cresol); low molecular weight (less than about 10 residues) polypeptides; proteins, such as serum albumin, gelatin, or immunoglobulins; hydrophilic polymers such as polyvinylpyrrolidone; amino acids such as glycine, glutamine, asparagine, histidine, arginine, or lysine; monosaccharides, di saccharides, and other carbohydrates, including glucose, mannose, or dextrins; chelating agents such as EDTA; sugars such as sucrose, mannitol, trehalose, or sorbitol; salt-forming counter-ions such as sodium; metal complexes (e.g., Zn-protein complexes); and / or non-ionic surfactants such as TWEEN™, PLURONICS™ or polyethylene glycol (PEG).

[0320] In a specific embodiment, pharmaceutical compositions comprise an anti-LY6K ADC disclosed herein, and optionally one or more additional prophylactic or therapeutic agents, in a pharmaceutically acceptable carrier. In a specific embodiment, pharmaceutical compositions comprise an anti-LY6K ADC disclosed herein, and optionally one or more additional prophylactic or therapeutic agents, in a pharmaceutically acceptable carrier. In certain embodiments, the ADC is the only active ingredient included in the pharmaceutical composition. Pharmaceutical compositions described herein can be useful in decreasing or blocking LY6K (e.g., human LY6K) activity and treating a condition, such as cancer. In certain embodiments, the present disclosure relates to a pharmaceutical composition of the present disclosure comprising an anti-LY6K ADC of the present disclosure for use as a medicament. In another embodiment, the present disclosurerelates to a pharmaceutical composition of the present disclosure for use in a method for the treatment of cancer.

[0321] Pharmaceutically acceptable carriers used in parenteral preparations include aqueous vehicles, nonaqueous vehicles, antimicrobial agents, isotonic agents, buffers, antioxidants, local anesthetics, suspending and dispersing agents, emulsifying agents, sequestering or chelating agents, and other pharmaceutically acceptable substances. Examples of aqueous vehicles include Sodium Chloride Injection, Ringer’s Injection, Isotonic Dextrose Injection, Sterile Water Injection, Dextrose and Lactated Ringer’s Injection. Nonaqueous parenteral vehicles include fixed oils of vegetable origin, cottonseed oil, corn oil, sesame oil, and peanut oil. Antimicrobial agents in bacteriostatic or fungistatic concentrations can be added to parenteral preparations packaged in multiple-dose containers which include phenols or cresols, mercurials, benzyl alcohol, chlorobutanol, methyl and propyl p-hydroxybenzoic acid esters, thimerosal, benzalkonium chloride, and benzethonium chloride. Isotonic agents include sodium chloride and dextrose. Buffers include phosphate and citrate. Antioxidants include sodium bisulfate. Local anesthetics include procaine hydrochloride. Suspending and dispersing agents include sodium carboxymethylcellulose, hydroxypropyl methylcellulose, and polyvinylpyrrolidone. Emulsifying agents include Polysorbate 80 (TWEEN® 80). A sequestering or chelating agent of metal ions includes EDTA. Pharmaceutical carriers also include ethyl alcohol, polyethylene glycol, and propylene glycol for water miscible vehicles; and sodium hydroxide, hydrochloric acid, citric acid, or lactic acid for pH adjustment.

[0322] A pharmaceutical composition can be formulated for any route of administration to a subject. Specific examples of routes of administration include intranasal, oral, pulmonary, transdermal, intradermal, and parenteral. Parenteral administration, characterized by either subcutaneous, intramuscular, or intravenous injection, is also contemplated herein. Injectables can be prepared in conventional forms, either as liquid solutions or suspensions, solid forms suitable for solution or suspension in liquid prior to injection, or as emulsions. The injectables, solutions, and emulsions also contain one or more excipients. Suitable excipients are, for example, water, saline, dextrose, glycerol, or ethanol. In addition, if desired, the pharmaceutical compositions to be administered can also contain minor amounts of non-toxic auxiliary substances such as wetting or emulsifying agents, pH buffering agents, stabilizers, solubility enhancers, and other such agents,such as for example, sodium acetate, sorbitan monolaurate, triethanolamine oleate, and cyclodextrins.

[0323] Preparations for parenteral administration of an ADC include sterile solutions ready for injection, sterile dry soluble products, such as lyophilized powders, ready to be combined with a solvent just prior to use, including hypodermic tablets, sterile suspensions ready for injection, sterile dry insoluble products ready to be combined with a vehicle just prior to use, and sterile emulsions. The solutions may be either aqueous or nonaqueous.

[0324] If administered intravenously, suitable carriers include physiological saline or phosphate buffered saline (PBS), and solutions containing thickening and solubilizing agents, such as glucose, polyethylene glycol, and polypropylene glycol, and mixtures thereof.

[0325] Topical mixtures comprising an ADC are prepared as described for the local and systemic administration. The resulting mixture can be a solution, suspension, emulsions, or the like and can be formulated as creams, gels, ointments, emulsions, solutions, elixirs, lotions, suspensions, tinctures, pastes, foams, aerosols, irrigations, sprays, suppositories, bandages, dermal patches, or any other formulations suitable for topical administration.

[0326] An anti-LY6K ADC disclosed herein can be formulated as an aerosol for topical application, such as by inhalation (see, e.g., U. S. Patent Nos. 4,044,126, 4,414,209, and 4,364,923, which describe aerosols for delivery of a steroid useful for treatment of inflammatory diseases, particularly asthma and are herein incorporated by reference in their entireties). These formulations for administration to the respiratory tract can be in the form of an aerosol or solution for a nebulizer, or as a microfine powder for insufflations, alone or in combination with an inert carrier such as lactose. In such a case, the particles of the formulation will, in certain embodiments, have diameters of less than 50 microns, in certain embodiments, less than 10 microns.

[0327] An anti-LY6K ADC disclosed herein can be formulated for local or topical application, such as for topical application to the skin and mucous membranes, such as in the eye, in the form of gels, creams, and lotions and for application to the eye or for intracistemal or intraspinal application. Topical administration is contemplated for transdermal delivery and also for administration to the eyes or mucosa, or for inhalation therapies. Nasal solutions of the ADC alone or in combination with other pharmaceutically acceptable excipients can also be administered.

[0328] Transdermal patches, including iontophoretic and electrophoretic devices, are well known to those of skill in the art, and can be used to administer an ADC. For example, such patchesare disclosed in U. S. Patent Nos. 6,267,983, 6,261,595, 6,256,533, 6,167,301, 6,024,975, 6,010715, 5,985,317, 5,983,134, 5,948,433, and 5,860,957, all of which are herein incorporated by reference in their entireties.

[0329] In certain embodiments, a pharmaceutical composition comprising an ADC described herein is a lyophilized powder, which can be reconstituted for administration as solutions, emulsions, and other mixtures. It may also be reconstituted and formulated as solids or gels. The lyophilized powder is prepared by dissolving an ADC described herein, or a pharmaceutically acceptable derivative thereof, in a suitable solvent. In certain embodiments, the lyophilized powder is sterile. The solvent may contain an excipient which improves the stability or other pharmacological component of the powder or reconstituted solution, prepared from the powder. Excipients that may be used include, but are not limited to, dextrose, sorbitol, fructose, com syrup, xylitol, glycerin, glucose, sucrose, or other suitable agent. The solvent may also contain a buffer, such as citrate, sodium or potassium phosphate, or other such buffer known to those of skill in the art, at, in certain embodiments, about neutral pH. Subsequent sterile fdtration of the solution followed by lyophilization under standard conditions known to those of skill in the art provides the desired formulation. In certain embodiments, the resulting solution will be apportioned into vials for lyophilization. Each vial will contain a single dosage or multiple dosages of the compound. The lyophilized powder can be stored under appropriate conditions, such as at about 4°C to room temperature. Reconstitution of this lyophilized powder with water for injection provides a formulation for use in parenteral administration. For reconstitution, the lyophilized powder is added to sterile water or other suitable carrier. The precise amount depends upon the selected compound. Such an amount can be empirically determined.

[0330] The anti-LY6K ADCs disclosed herein and other compositions provided herein can also be formulated to be targeted to a particular tissue, receptor, or other area of the body of the subject to be treated. Many such targeting methods are well known to those of skill in the art. All such targeting methods are contemplated herein for use in the instant compositions. For nonlimiting examples of targeting methods, see, e.g., U. S. Patent Nos. 6,316,652, 6,274,552, 6,271,359, 6,253,872, 6,139,865, 6,131,570, 6,120,751, 6,071,495, 6,060,082, 6,048,736, 6,039,975, 6,004,534, 5,985,307, 5,972,366, 5,900,252, 5,840,674, 5,759,542, and 5,709,874, all of which are herein incorporated by reference in their entireties. In a specific embodiment, an ADC described herein is targeted to a tumor.

[0331] The compositions to be used for in vivo administration can be sterile. This is readily accomplished by filtration through, e.g., sterile filtration membranes.Methods of Use and Uses

[0332] In another aspect, the instant disclosure provides a method of treating a subject using the anti-LY6K ADCs disclosed herein. Any disease or disorder in a subject that is characterized by overexpression of LY6K (e.g., human LY6K) can be treated using the anti-LY6K ADCs disclosed herein. In certain embodiments, the disease or disorder is resistant to a checkpoint targeting agent (e.g., an antagonist anti-CTLA-4 antibody, an antagonist anti-PD-Ll antibody, an antagonist anti-PD-L2 antibody, or an antagonist anti-PD-1 antibody). In certain embodiments, the disease or disorder is recurrent after treatment with a checkpoint targeting agent (e.g., an antagonist anti-CTLA-4 antibody, an antagonist anti-PD-Ll antibody, an antagonist anti-PD-L2 antibody, or an antagonist anti-PD-1 antibody).

[0333] In certain embodiments, the instant disclosure provides a method of treating cancer in a subject, the method comprising administering to the subject an effective amount of the ADC or pharmaceutical composition as disclosed herein.

[0334] Cancers that can be treated with the anti-LY 6K ADCs or pharmaceutical compositions disclosed herein include, without limitation, a solid tumor, a hematological cancer (e.g., leukemia, lymphoma, myeloma, e.g., multiple myeloma), and a metastatic lesion. In certain embodiments, the cancer is a solid tumor. Examples of solid tumors include malignancies, e.g., sarcomas and carcinomas, e.g., adenocarcinomas of the various organ systems, such as those affecting the lung, breast, ovarian, lymphoid, gastrointestinal (e.g., colon), anal, genitals and genitourinary tract (e.g., renal, urothelial, bladder cells, prostate), pharynx, CNS (e.g., brain, neural or glial cells), head and neck, skin (e.g., melanoma), and pancreas, as well as adenocarcinomas which include malignancies such as colon cancers, rectal cancer, renal-cell carcinoma, liver cancer, lung cancer (e.g., non-small cell lung cancer or small cell lung cancer), cancer of the small intestine, and cancer of the esophagus. The cancer may be at an early, intermediate, late stage, or metastatic cancer. In certain embodiments, the cancer is resistant to a checkpoint targeting agent (e.g., an antagonist anti-CTLA-4 antibody, an antagonist anti-PD-Ll antibody, an antagonist anti-PD-L2 antibody, or an antagonist anti-PD-1 antibody). In certain embodiments, the cancer is recurrent after treatment with a checkpoint targeting agent (e.g., an antagonist anti-CTLA-4 antibody, an antagonist anti-PD-Ll antibody, an antagonist anti-PD-L2 antibody, or an antagonist anti-PD-1 antibody).

[0335] In certain embodiments, the cancer is chosen from lung cancer (e.g., lung adenocarcinoma or non-small cell lung cancer (NSCLC) (e.g., NSCLC with squamous and / or non-squamous histology, or NSCLC adenocarcinoma)), melanoma e.g., an advanced melanoma), renal cancer (e.g., a renal cell carcinoma), liver cancer (e.g., hepatocellular carcinoma), myeloma (e.g., a multiple myeloma), a prostate cancer, a breast cancer (e.g., a breast cancer that does not express one, two or all of estrogen receptor, progesterone receptor, or Her2 / neu, e.g., a triple negative breast cancer), an ovarian cancer, a colorectal cancer, a pancreatic cancer, a head and neck cancer (e.g., head and neck squamous cell carcinoma (HNSCC)), anal cancer, gastroesophageal cancer (e.g., esophageal squamous cell carcinoma), mesothelioma, nasopharyngeal cancer, thyroid cancer, cervical cancer, epithelial cancer, peritoneal cancer, or a lymphoproliferative disease (e.g., a post-transplant lymphoproliferative disease).

[0336] In certain embodiments, the cancer is a hematological cancer, for example, a leukemia, a lymphoma, or a myeloma. In certain embodiments, the cancer is a leukemia, for example, acute lymphoblastic leukemia (ALL), acute myelogenous leukemia (AML), acute myeloblastic leukemia (AML), chronic lymphocytic leukemia (CLL), chronic myelogenous leukemia (CML), chronic myeloid leukemia (CML), chronic myelomonocytic leukemia (CMML), chronic lymphocytic leukemia (CLL), or hairy cell leukemia. In certain embodiments, the cancer is a lymphoma, for example, B cell lymphoma, diffuse large B-cell lymphoma (DLBCL), activated B-cell like (ABC) diffuse large B cell lymphoma, germinal center B cell (GCB) diffuse large B cell lymphoma, mantle cell lymphoma, Hodgkin lymphoma, non-Hodgkin lymphoma, relapsed nonHodgkin lymphoma, refractory non-Hodgkin lymphoma, recurrent follicular non-Hodgkin lymphoma, Burkitt lymphoma, small lymphocytic lymphoma, follicular lymphoma, lymphoplasmacytic lymphoma, or extranodal marginal zone lymphoma. In certain embodiments the cancer is a myeloma, for example, multiple myeloma.

[0337] In another embodiment, the cancer is chosen from a carcinoma (e.g., advanced or metastatic carcinoma), melanoma or a lung carcinoma, e.g., a non-small cell lung carcinoma.

[0338] In certain embodiments, the cancer is a lung cancer, e.g., a lung adenocarcinoma, non-small cell lung cancer, or small cell lung cancer.

[0339] In certain embodiments, the cancer is a melanoma, e.g., an advanced melanoma. In certain embodiments, the cancer is an advanced or unresectable melanoma that does not respond to other therapies. In other embodiments, the cancer is a melanoma with a BRAF mutation (e.g., aBRAF V600 mutation). In yet other embodiments, the anti-LY6K ADC or pharmaceutical composition disclosed herein is administered after treatment with an anti-CTLA-4 antibody (e.g., ipilimumab) with or without a BRAF inhibitor (e.g., vemurafenib or dabrafenib).

[0340] In another embodiment, the cancer is a hepatocarcinoma, e.g., an advanced hepatocarcinoma, with or without a viral infection, e.g., a chronic viral hepatitis.

[0341] In another embodiment, the cancer is a prostate cancer, e.g., an advanced prostate cancer.

[0342] In yet another embodiment, the cancer is a myeloma, e.g., multiple myeloma.

[0343] In yet another embodiment, the cancer is a renal cancer, e.g., a renal cell carcinoma (RCC) (e.g., a metastatic RCC, clear cell renal cell carcinoma (CCRCC) or kidney papillary cell carcinoma).

[0344] In yet another embodiment, the cancer is chosen from a lung cancer, a melanoma, a renal cancer, a breast cancer, a colorectal cancer, a leukemia, or a metastatic lesion of the cancer.

[0345] In certain embodiments, these methods further comprise administering an additional therapeutic agent to the subject. In certain embodiments, the additional therapeutic agent is a chemotherapeutic, a radiotherapeutic, or a checkpoint targeting agent. In certain embodiments, the chemotherapeutic agent is a hypomethylating agent (e.g., azacitidine). In certain embodiments, the chemotherapeutic agent is a DNA damage-inducing agent (e.g., gemcitabine). In certain embodiments, the checkpoint targeting agent is selected from the group consisting of an antagonist anti-CTLA-4 antibody, an antagonist anti-PD-Ll antibody, an antagonist anti-PD-L2 antibody, an antagonist anti-PD-1 antibody, an antagonist anti-TIM-3 antibody, an antagonist anti -LAG-3 antibody, an antagonist anti-VISTA antibody, an antagonist anti-CD96 antibody, an antagonist anti-CEACAMl antibody, an agonist anti-CD137 antibody, an agonist anti-GITR antibody, and an agonist anti-OX40 antibody. In certain embodiments, the checkpoint targeting agent is selected from the group consisting of an antagonist anti-CTLA-4 antibody, an antagonist anti-PD-Ll antibody, an antagonist anti-PD-L2 antibody, and an antagonist anti-PD-1 antibody, wherein the LY6K (e.g., human LY6K) ADCs or pharmaceutical compositions disclosed herein synergize with the checkpoint targeting agent.

[0346] In certain embodiments, the present disclosure relates to an ADC and / or pharmaceutical composition of the present disclosure for use in a method of the present disclosure, wherein the method further comprises administering an additional therapeutic agent to the subject.In certain embodiments, the present disclosure relates to (a) an ADC and / or pharmaceutical composition of the present disclosure and (b) an additional therapeutic agent for use as a medicament. In certain embodiments, the present disclosure relates to (a) an ADC and / or pharmaceutical composition of the present disclosure and (b) an additional therapeutic agent for use in a method for the treatment of cancer. In a further embodiment, the present disclosure relates to a pharmaceutical composition, kit or kit-of-parts comprising (a) an ADC and / or pharmaceutical composition of the present disclosure and (b) an additional therapeutic agent. In certain embodiments, the additional therapeutic agent is a chemotherapeutic, a radiotherapeutic, or a checkpoint targeting agent.

[0347] In certain embodiments, an anti-PD-1 antibody is used in methods disclosed herein. In certain embodiments, the anti-PD-1 antibody is nivolumab, also known as BMS-936558 or MDX1106, developed by Bristol-Myers Squibb. In certain embodiments, the anti-PD-1 antibody is pembrolizumab, also known as lambrolizumab or MK-3475, developed by Merck & Co. In certain embodiments, the anti-PD-1 antibody is pidilizumab, also known as CT-011, developed by CureTech. In certain embodiments, the anti-PD-1 antibody is MEDI0680, also known as AMP-514, developed by Medlmmune. In certain embodiments, the anti-PD-1 antibody is PDR001 developed by Novartis Pharmaceuticals. In certain embodiments, the anti-PD-1 antibody is REGN2810 developed by Regeneron Pharmaceuticals. In certain embodiments, the anti-PD-1 antibody is PF-06801591 developed by Pfizer. In certain embodiments, the anti-PD-1 antibody is BGB-A317 developed by BeiGene. In certain embodiments, the anti-PD-1 antibody is TSR-042 developed by AnaptysBio and Tesaro. In certain embodiments, the anti-PD-1 antibody is SHR-1210 developed by Hengrui.

[0348] Further non-limiting examples of anti-PD-1 antibodies that may be used in treatment methods disclosed herein are disclosed in the following patents and patent applications, all of which are herein incorporated by reference in their entireties for all purposes: U. S. Patent No.6,808,710; U. S. PatentNo. 7,332,582; U. S. PatentNo. 7,488,802; U. S. PatentNo. 8,008,449; U. S. Patent No. 8,114,845; U. S. Patent No. 8,168,757; U. S. Patent No. 8,354,509; U. S. Patent No.8,686,119; U. S. PatentNo. 8,735,553; U. S. PatentNo. 8,747,847; U. S. PatentNo. 8,779,105; U. S. Patent No. 8,927,697; U. S. Patent No. 8,993,731; U. S. Patent No. 9,102,727; U. S. Patent No.9,205,148; U. S. Publication No. US 2013 / 0202623 Al; U. S. Publication No. US 2013 / 0291136 Al; U. S. Publication No. US 2014 / 0044738 Al; U. S. Publication No. US 2014 / 0356363 Al; U. S.Publication No. US 2016 / 0075783 Al; PCT Publication No. WO 2013 / 033091 Al; PCT Publication No. WO 2015 / 036394 Al; PCT Publication No. WO 2014 / 179664 A2; PCT Publication No. WO 2014 / 209804 Al; PCT Publication No. WO 2014 / 206107 Al; PCT Publication No. WO 2015 / 058573 Al; PCT Publication No. WO 2015 / 085847 Al; PCT Publication No. WO 2015 / 200119 Al; PCT Publication No. WO 2016 / 015685 Al; and PCT Publication No. WO 2016 / 020856 AL

[0349] In certain embodiments, an anti-PD-Ll antibody is used in methods disclosed herein. In certain embodiments, the anti-PD-Ll antibody is atezolizumab developed by Genentech. In certain embodiments, the anti-PD-Ll antibody is durvalumab developed by AstraZeneca, Celgene, and Medlmmune. In certain embodiments, the anti-PD-Ll antibody is avelumab, also known as MSB0010718C, developed by Merck Serono and Pfizer. In certain embodiments, the anti-PD-Ll antibody is MDX-1105 developed by Bristol-Myers Squibb. In certain embodiments, the anti-PD-Ll antibody is AMP-224 developed by Amplimmune and GSK.

[0350] Non-limiting examples of anti-PD-L 1 antibodies that may be used in treatment methods disclosed herein are disclosed in the following patents and patent applications, all of which are herein incorporated by reference in their entireties for all purposes: U. S. Patent No. 7,943,743; U. S. Patent No. 8,168,179; U. S. Patent No. 8,217,149; U. S. Patent No. 8,552,154; U. S. Patent No.8,779,108; U. S. Patent No. 8,981,063; U. S. Patent No. 9,175,082; U. S. Publication No. US 2010 / 0203056 Al; U. S. Publication No. US 2003 / 0232323 Al; U. S Publication No. US 2013 / 0323249 Al; U. S. Publication No. US 2014 / 0341917 Al; U. S Publication No. US 2014 / 0044738 Al; U. S. Publication No. US 2015 / 0203580 Al; U. S Publication No. US 2015 / 0225483 Al; U. S. Publication No. US 2015 / 0346208 Al; U. S Publication No. US 2015 / 0355184 Al; PCT Publication No. WO 2014 / 100079 Al; PCT Publication No. WO 2014 / 022758 Al; PCT Publication No. WO 2014 / 055897 A2; PCT Publication No. WO 2015 / 061668 Al; PCT Publication No. WO 2015 / 109124 Al; PCT Publication No. WO 2015 / 195163 Al; PCT Publication No. WO 2016 / 000619 Al; and PCT Publication No. WO 2016 / 030350 Al.

[0351] In certain embodiments, an anti-CTLA-4 antibody is used in methods disclosed herein. In certain embodiments, the anti-CTLA-4 antibody is ipilimumab developed by Bristol-Myers Squibb.

[0352] In certain embodiments, an anti-LY6K ADC or pharmaceutical composition disclosed herein is administered to a subject in combination with a compound that targets an immunomodulatory enzyme(s) such as IDO (indoleamine-(2,3)-dioxygenase) and / or TDO (tryptophan 2,3-dioxygenase). Therefore, in certain embodiments, the additional therapeutic agent is a compound that targets an immunomodulatory enzyme(s), such as an inhibitor of indoleamine-(2, 3 )-di oxygenase (IDO). In certain embodiments, such compound is selected from the group consisting of epacadostat (Incyte Corp; see, e.g., WO 2010 / 005958 which is herein incorporated by reference in its entirety), F001287 (Flexus Biosciences / Bristol-Myers Squibb), indoximod (NewLink Genetics), and NLG919 (NewLink Genetics). In certain embodiments, the compound is epacadostat. In another embodiment, the compound is F001287. In another embodiment, the compound is indoximod. In another embodiment, the compound is NLG919. In a specific embodiment, an anti-LY6K ADC or pharmaceutical composition disclosed herein is administered to a subject in combination with an IDO inhibitor for treating cancer. The IDO inhibitor as described herein for use in treating cancer is present in a solid dosage form of a pharmaceutical composition such as a tablet, a pill, or a capsule, wherein the pharmaceutical composition includes an IDO inhibitor and a pharmaceutically acceptable excipient. As such, the ADC or pharmaceutical composition as described herein and the IDO inhibitor as described herein can be administered separately, sequentially, or concurrently as separate dosage forms. In certain embodiments, the ADC or pharmaceutical composition is administered parenterally, and the IDO inhibitor is administered orally. In certain embodiments, the inhibitor is selected from the group consisting of epacadostat (Incyte Corporation), F001287 (Flexus Biosciences / Bristol-Myers Squibb), indoximod (NewLink Genetics), and NLG919 (NewLink Genetics). Epacadostat has been described in PCT Publication No. WO 2010 / 005958, which is herein incorporated by reference in its entirety for all purposes. In certain embodiments, the inhibitor is epacadostat. In another embodiment, the inhibitor is F001287. In another embodiment, the inhibitor is indoximod. In another embodiment, the inhibitor is NLG919.

[0353] In certain embodiments, an anti-LY6K ADC or pharmaceutical composition disclosed herein is administered to a subject in combination with a vaccine. The vaccine can be, e.g., a peptide vaccine, a DNA vaccine, or an RNA vaccine.

[0354] In certain embodiments, an anti-LY6K ADC or pharmaceutical composition disclosed herein is administered to a subject in combination with an adjuvant. Various adjuvants can be useddepending on the treatment context. Non-limiting examples of appropriate adjuvants include, but not limited to, Complete Freund’s Adjuvant (CFA), Incomplete Freund’s Adjuvant (IF A), montanide ISA (incomplete Seppic adjuvant), the Ribi adjuvant system (RAS), Titer Max, muramyl peptides, Syntex Adjuvant Formulation (SAF), alum (aluminum hydroxide and / or aluminum phosphate), aluminum salt adjuvants, Gerbu® adjuvants, nitrocellulose absorbed antigen, encapsulated or entrapped antigen, 3 De-O-acylated monophosphoryl lipid A (3 D-MPL), immunostimulatory oligonucleotides, toll-like receptor (TLR) ligands, mannan-binding lectin (MBL) ligands, STING agonists, immuno-stimulating complexes such as saponins, Quil A, QS-21, QS-7, ISCOMATRIX, and others. Other adjuvants include CpG oligonucleotides and double stranded RNA molecules, such as poly(A) and poly(U). Combinations of the above adjuvants may also be used. See, e.g., U. S. Patent Nos. 6,645,495; 7,029,678; and 7,858,589, all of which are incorporated herein by reference in their entireties. In certain embodiments, the adjuvant used herein is QS-21 STIMULON.

[0355] In certain embodiments, an anti-LY6K ADC or pharmaceutical composition disclosed herein is administered to a subject in combination with an additional therapeutic agent comprising a TCR. In certain embodiments, the additional therapeutic agent is a soluble TCR. In certain embodiments, the additional therapeutic agent is a cell expressing a TCR. Therefore, in certain embodiments, the present disclosure relates to an anti-LY6K ADC or pharmaceutical composition of the present disclosure in combination with an additional therapeutic agent comprising a TCR for use as a medicament and / or for use in a method for the treatment of cancer.

[0356] In certain embodiments, an anti-LY6K ADC or pharmaceutical composition disclosed herein is administered to a subject in combination with a cell expressing a chimeric antigen receptor (CAR). In certain embodiments, the cell is a T cell.

[0357] In certain embodiments, an anti-LY6K ADC or pharmaceutical composition disclosed herein is administered to a subject in combination with a TCR mimic antibody. In certain embodiments, the TCR mimic antibody is an antibody that specifically binds to a peptide-MHC complex. For non -limiting examples of TCR mimic antibodies, see, e.g., U. S. Patent No. 9,074,000 and U. S. Publication Nos. US 2009 / 0304679 Al and US 2014 / 0134191 Al, all of which are incorporated herein by reference in their entireties.

[0358] In certain embodiments, an anti-LY6K ADC or pharmaceutical composition disclosed herein is administered to a subject in combination with a bispecific T-cell engager (BiTE) (e.g., asdescribed in W02005061547A2, which is incorporated by reference herein in its entirety) and / or a dual-affinity re-targeting antibody (DART) (e.g., as described in WO2012162067A2, which is incorporated by reference herein in its entirety). In certain embodiments, the BiTE and / or DART specifically binds to a tumor-associated antigen (e.g., a polypeptide overexpressed in a tumor, a polypeptide derived from an oncovirus, a polypeptide comprising a post-translational modification specific to a tumor, a polypeptide specifically mutated in a tumor) and a molecule on an effector cell (e.g., CD3 or CD16). In certain embodiments, the tumor-associated antigen is EGFR (e.g., human EGFR), optionally wherein the BiTE and / or DART comprises the VH and VL sequences of cetuximab. In certain embodiments, the tumor-associated antigen is Her2 (e.g., human Her2), optionally wherein the BiTE and / or DART comprises the VH and VL sequences of trastuzumab. In certain embodiments, the tumor-associated antigen is CD20 (e.g., human CD20).

[0359] The anti-LY6K ADC or pharmaceutical composition and the additional therapeutic agent (e.g., chemotherapeutic, radiotherapeutic, checkpoint targeting agent, IDO inhibitor, vaccine, adjuvant, a soluble TCR, a cell expressing a TCR, a cell expressing a chimeric antigen receptor, and / or a TCR mimic antibody) can be administered separately, sequentially, or concurrently as separate dosage forms. In certain embodiments, an anti-LY6K ADC or pharmaceutical composition provided herein is administered parenterally, and an IDO inhibitor is administered orally.

[0360] An anti-LY6K ADC or pharmaceutical composition described herein may be delivered to a subject by a variety of routes. These include, but are not limited to, parenteral, intranasal, intratracheal, oral, intradermal, topical, intramuscular, intraperitoneal, transdermal, intravenous, intratumoral, conjunctival, intra-arterial, and subcutaneous routes. Pulmonary administration can also be employed, e.g., by use of an inhaler or nebulizer, and formulation with an aerosolizing agent for use as a spray. In certain embodiments, the anti-LY6K ADC or pharmaceutical composition described herein is delivered subcutaneously or intravenously. In certain embodiments, the anti-LY6K ADC or pharmaceutical composition described herein is delivered intra-arterially. In certain embodiments, the anti-LY6K ADC or pharmaceutical composition described herein is delivered intratum orally. In certain embodiments, the anti-LY6K ADC or pharmaceutical composition described herein is delivered into a tumor draining lymph node.

[0361] The amount of an ADC or composition which will be effective in the treatment and / or prevention of a condition will depend on the nature of the disease and can be determined by standard clinical techniques.

[0362] The precise dose to be employed in a composition will also depend on the route of administration, and the seriousness of the infection or disease caused by it and should be decided according to the judgment of the practitioner and each subject’s circumstances. For example, effective doses may also vary depending upon means of administration, target site, physiological state of the patient (including age, body weight, and health), whether the patient is human or an animal, other medications administered, or whether treatment is prophylactic or therapeutic. Usually, the patient is a human, but non-human mammals, including transgenic mammals, can also be treated. Treatment dosages are optimally titrated to optimize safety and efficacy.

[0363] An anti-LY6K ADC described herein can be used for prognostic, diagnostic, monitoring, and screening applications, including iti vitro and in vivo applications well known and standard to the skilled artisan and based on the present description. Prognostic, diagnostic, monitoring, and screening assays and kits for in vitro assessment and evaluation of immune system status, immune response, and / or cancer (e.g., tumorigenesis) may be utilized to predict, diagnose, and monitor to evaluate patient samples, including those known to have or suspected of having an immune system-dysfunction or cancer, or with regard to an anticipated or desired immune system response, antigen response, or vaccine response. The assessment and evaluation of immune system status, immune response, and / or cancer are also useful in determining the suitability of a patient for a clinical trial of a drug or for the administration of a particular chemotherapeutic agent, a radiotherapeutic agent, or an antibody, including combinations thereof, versus a different agent or antibody. This type of prognostic and diagnostic monitoring and assessment is already in practice utilizing antibodies against the HER2 protein in breast cancer (HercepTest™, Dako) where the assay is also used to evaluate patients for antibody therapy using Herceptin®. In vivo applications include directed cell therapy and immune system modulation and radio imaging of immune responses. Therefore, in certain embodiments, the present disclosure relates to an anti-LY6K ADC and / or pharmaceutical composition of the present disclosure for use as a diagnostic. In certain embodiments, the present disclosure relates to an anti-LY6K ADC and / or pharmaceutical composition of the present disclosure for use in a method for the prediction, diagnosis, and / or monitoring of a subject having or suspected to have an immune system-dysfunction, cancer, and / orwith regard to an anticipated or desired immune system response, antigen response, or vaccine response. In another embodiment, the present disclosure relates to the use of an anti-LY6K ADC of the disclosure, for predicting, diagnosing, and / or monitoring of a subject having or suspected to have an immune system-dysfunction, cancer, and / or with regard to an anticipated or desired immune system response, antigen response, or vaccine response by assaying and / or detecting human LY6K protein levels in a biological sample of the subject in vitro.Polynucleotides, Vectors, and Methods of Producing Antibodies

[0364] Also described herein are polynucleotides comprising a nucleotide sequence encoding an antibody, or a portion thereof, described herein or a fragment thereof (e.g., a VL and / or VH; and a light chain and / or heavy chain) that specifically binds to a LY6K (e.g., human LY6K) antigen, and vectors, e.g., vectors comprising such polynucleotides for recombinant expression in host cells (e.g., E. coli and mammalian cells). Described herein are polynucleotides comprising nucleotide sequences encoding a heavy and / or light chain of any of the antibodies described herein, as well as vectors comprising such polynucleotide sequences, e.g., expression vectors for their efficient expression in host cells, e.g., mammalian cells. In certain embodiments, a nucleic acid molecule(s) encoding an antibody described herein is isolated or purified.

[0365] In particular aspects, described herein are polynucleotides comprising nucleotide sequences encoding antibodies, which specifically bind to a LY6K (e.g., human LY6K) polypeptide and comprise an amino acid sequence as described herein, as well as antibodies which compete with such antibodies for binding to a LY6K (e.g., human LY6K) polypeptide (e.g., in a dose-dependent manner), or which binds to the same epitope as that of such antibodies.

[0366] In certain aspects, described herein are polynucleotides comprising a nucleotide sequence encoding the light chain or heavy chain of an antibody described herein. The polynucleotides can comprise nucleotide sequences encoding a light chain comprising the VL FRs and CDRs of antibodies described herein (see, e.g., Tables 2 and 4) or nucleotide sequences encoding a heavy chain comprising the VH FRs and CDRs of antibodies described herein (see, e.g., Tables 1 and 3). In certain embodiments, a polynucleotide encodes a VH, VL, heavy chain, and / or light chain of an antibody described herein. In another embodiment, a polynucleotide encodes the first VH and the first VL of an antibody described herein. In another embodiment, a polynucleotide encodes the second VH and the second VL of an antibody described herein. In another embodiment, a polynucleotide encodes the first heavy chain and the first light chain of anantibody described herein. In another embodiment, a polynucleotide encodes the second heavy chain and the second light chain of an antibody described herein. In another embodiment, a polynucleotide encodes the VH and / or the VL, or the heavy chain and / or the light chain, of an antibody described herein.

[0367] Also described herein are polynucleotides encoding an anti-LY6K antibody that are optimized, e.g., by codon / RNA optimization, replacement with heterologous signal sequences, and elimination of mRNA instability elements. Methods to generate optimized nucleic acids encoding an anti-LY6K antibody or a fragment thereof (e.g., light chain, heavy chain, VH domain, or VL domain) for recombinant expression by introducing codon changes and / or eliminating inhibitory regions in the mRNA can be carried out by adapting the optimization methods described in, e.g., U. S. Patent Nos. 5,965,726; 6,174,666; 6,291,664; 6,414,132; and 6,794,498, accordingly, all of which are herein incorporated by reference in their entireties. For example, potential splice sites and instability elements (e.g., A / T or A / U rich elements) within the RNA can be mutated without altering the amino acids encoded by the nucleic acid sequences to increase stability of the RNA for recombinant expression. The alterations utilize the degeneracy of the genetic code, e.g., using an alternative codon for an identical amino acid. In certain embodiments, it can be desirable to alter one or more codons to encode a conservative mutation, e.g., a similar amino acid with similar chemical structure and properties and / or function as the original amino acid. Such methods can increase expression of an anti-LY6K antibody or fragment thereof by at least 1 fold, 2 fold, 3 fold, 4 fold, 5 fold, 10 fold, 20 fold, 30 fold, 40 fold, 50 fold, 60 fold, 70 fold, 80 fold, 90 fold, or 100 fold or more relative to the expression of an anti-LY6K antibody encoded by polynucleotides that have not been optimized.

[0368] In certain embodiments, an optimized polynucleotide sequence encoding an anti-LY6K antibody described herein or a fragment thereof (e.g., VL domain and / or VH domain) can hybridize to an antisense (e.g., complementary) polynucleotide of an unoptimized polynucleotide sequence encoding an anti-LY6K antibody described herein or a fragment thereof (e.g., VL domain and / or VH domain). In specific embodiments, an optimized nucleotide sequence encoding an anti-LY6K antibody described herein or a fragment thereof hybridizes under high stringency conditions to antisense polynucleotide of an unoptimized polynucleotide sequence encoding an anti-LY6K antibody described herein or a fragment thereof. In a specific embodiment, an optimized nucleotide sequence encoding an anti-LY6K antibody described herein or a fragmentthereof hybridizes under high stringency, intermediate or lower stringency hybridization conditions to an antisense polynucleotide of an unoptimized nucleotide sequence encoding an anti-LY6K antibody described herein or a fragment thereof. Information regarding hybridization conditions has been described, see, e.g., U. S. Patent Application Publication No. US 2005 / 0048549 (e.g., paragraphs 72-73), which is herein incorporated by reference in its entirety.

[0369] The polynucleotides can be obtained, and the nucleotide sequence of the polynucleotides determined, by any method known in the art. Nucleotide sequences encoding antibodies described herein, e.g., antibodies described in Tables 1-4, and modified versions of these antibodies can be determined using methods known in the art, i.e., nucleotide codons known to encode particular amino acids are assembled in such a way to generate a nucleic acid that encodes the antibody. Such a polynucleotide encoding the antibody can be assembled from chemically synthesized oligonucleotides (e.g., as described in Kutmeier G et al., (1994), BioTechniques 17: 242-6, herein incorporated by reference in its entirety), which, briefly, involves the synthesis of overlapping oligonucleotides containing portions of the sequence encoding the antibody, annealing and ligating of those oligonucleotides, and then amplification of the ligated oligonucleotides by PCR.[00370J Alternatively, a polynucleotide encoding an antigen-binding region of an antibody described herein can be generated from nucleic acid from a suitable source (e.g., a hybridoma) using methods well known in the art (e.g., PCR and other molecular cloning methods). For example, PCR amplification using synthetic primers hybridizable to the 3’ and 5’ ends of a known sequence can be performed using genomic DNA obtained from hybridoma cells producing the antibody of interest. Such PCR amplification methods can be used to obtain nucleic acids comprising the sequence encoding the light chain and / or heavy chain of an antibody. Such PCR amplification methods can be used to obtain nucleic acids comprising the sequence encoding the variable light chain region and / or the variable heavy chain region of an antibody. The amplified nucleic acids can be cloned into vectors for expression in host cells and for further cloning.

[0371] If a clone containing a nucleic acid encoding a particular antigen-binding region or antibody is not available, but the sequence of the antigen-binding region or antibody molecule is known, a nucleic acid encoding the immunoglobulin can be chemically synthesized or obtained from a suitable source (e.g., an antibody cDNA library or a cDNA library generated from, or nucleic acid, preferably poly A+ RNA, isolated from, any tissue or cells expressing the antibody,such as hybridoma cells selected to express an antibody described herein) by PCR amplification using synthetic primers hybridizable to the 3’ and 5’ ends of the sequence, or by cloning using an oligonucleotide probe specific for the particular gene sequence to identify, e.g., a cDNA clone from a cDNA library that encodes the antibody. Amplified nucleic acids generated by PCR can then be cloned into replicable cloning vectors using any method well known in the art.

[0372] DNA encoding anti-LY6K (e.g., human LY6K) antibodies described herein can be readily isolated and sequenced using conventional procedures (e.g., by using oligonucleotide probes that are capable of binding specifically to genes encoding the heavy and light chains of the anti-LY6K (e.g., human LY6K) antibodies). Hybridoma cells can serve as a source of such DNA. Once isolated, the DNA can be placed into expression vectors, which are then transfected into host cells such as E. coli cells, simian COS cells, Chinese hamster ovary (CHO) cells (e.g., CHO cells from the CHO GS System™ (Lonza)), or myeloma cells that do not otherwise produce immunoglobulin protein, to obtain the synthesis of anti-LY6K antibodies in the recombinant host cells.

[0373] To generate whole antibodies or antigen-binding regions, PCR primers including VH or VL nucleotide sequences, a restriction site, and a flanking sequence to protect the restriction site can be used to amplify the VH or VL sequences in scFv clones. Utilizing cloning techniques known to those of skill in the art, the PCR amplified VH domains can be cloned into vectors expressing a heavy chain constant region, e.g., the human gamma 1 or human gamma 4 constant region, and the PCR amplified VL domains can be cloned into vectors expressing a light chain constant region, e.g., human kappa or lambda constant regions. In certain embodiments, the vectors for expressing the VH or VL domains comprise an EF-la promoter, a secretion signal, a cloning site for the variable region, constant regions, and a selection marker such as neomycin. The VH and VL domains can also be cloned into one vector expressing the necessary constant regions. The heavy chain conversion vectors and light chain conversion vectors are then co-transfected into cell lines to generate stable or transient cell lines that express full-length antibodies, e.g., IgG, using techniques known to those of skill in the art.

[0374] The DNA also can be modified, for example, by substituting the coding sequence for human heavy and light chain constant regions in place of the murine sequences, or by covalently joining to the immunoglobulin coding sequence all or part of the coding sequence for a nonimmunoglobulin polypeptide.

[0375] Also described are polynucleotides that hybridize under high stringency, intermediate or lower stringency hybridization conditions to polynucleotides that encode an antibody described herein. In specific embodiments, polynucleotides described herein hybridize under high stringency, intermediate or lower stringency hybridization conditions to polynucleotides encoding a VH domain and / or VL domain described herein.

[0376] Hybridization conditions have been described in the art and are known to one of skill in the art. For example, hybridization under stringent conditions can involve hybridization to filterbound DNA in 6x sodium chloride / sodium citrate (SSC) at about 45°C followed by one or more washes in 0.2xSSC / 0.1% SDS at about 50-65°C; hybridization under highly stringent conditions can involve hybridization to filter-bound nucleic acid in 6xSSC at about 45°C followed by one or more washes in 0.1xSSC / 0.2% SDS at about 68°C. Hybridization under other stringent hybridization conditions are known to those of skill in the art and have been described, see, e.g., Ausubel FM et al., eds., (1989) Current Protocols in Molecular Biology, Vol. I, Green Publishing Associates, Inc. and John Wiley & Sons, Inc., New York at pages 6.3.1-6.3.6 and 2.10.3, which is herein incorporated by reference in its entirety.

[0377] In certain aspects, described herein are cells (e.g., host cells) expressing (e.g., recombinantly) antibodies described herein which specifically bind to LY6K (e.g., human LY6K), and related polynucleotides and expression vectors. Described herein are vectors (e.g., expression vectors) comprising polynucleotides comprising nucleotide sequences encoding anti-LY6K antibodies or a fragment for recombinant expression in host cells, preferably in mammalian cells (e.g., CHO cells). Also described herein are host cells comprising such vectors for recombinantly expressing anti-LY6K antibodies described herein (e.g., human or humanized antibody). In a particular aspect, described herein are methods for producing an antibody described herein, comprising expressing the antibody from a host cell.

[0378] Recombinant expression of an antibody described herein (e.g., a full-length antigenbinding region or antibody or heavy and / or light chain of an antibody described herein) that specifically binds to LY6K (e.g., human LY6K) generally involves construction of an expression vector containing a polynucleotide that encodes the antibody. Once a polynucleotide encoding an antibody molecule, heavy and / or light chain of an antibody, or a fragment thereof (e.g., heavy and / or light chain variable regions) described herein has been obtained, the vector for the production of the antibody molecule can be produced by recombinant DNA technology usingtechniques well known in the art. Thus, methods for preparing a protein by expressing a polynucleotide containing an antibody or antibody fragment (e.g., light chain or heavy chain) encoding nucleotide sequence are described herein. Methods which are well known to those skilled in the art can be used to construct expression vectors containing an antibody or antibody fragment (e.g., light chain or heavy chain) coding sequences and appropriate transcriptional and translational control signals. These methods include, for example, in vitro recombinant DNA techniques, synthetic techniques, and in vivo genetic recombination. Also described are replicable vectors comprising a nucleotide sequence encoding containing an antibody molecule described herein, a heavy or light chain of an antibody, a heavy or light chain variable region of an antibody or a fragment thereof, or a heavy or light chain CDR, operably linked to a promoter. Such vectors can, for example, include the nucleotide sequence encoding the constant region of the antibody molecule (see, e.g., International Publication Nos. WO 86 / 05807 and WO 89 / 01036; and U. S. Patent No. 5,122,464, which are herein incorporated by reference in their entireties) and variable regions of the antibody can be cloned into such a vector for expression of the entire heavy, the entire light chain, or both the entire heavy and light chains.

[0379] In certain embodiments, a vector comprises a polynucleotide encoding a VH, VL, heavy chain, and / or light chain of an antibody described herein. In another embodiment, a vector comprises a polynucleotide encoding the VH and the VL of an antibody described herein. In another embodiment, a vector comprises a polynucleotide encoding the heavy chain and the light chain of an antibody described herein.

[0380] An expression vector can be transferred to a cell (e.g., host cell) by conventional techniques and the resulting cells can then be cultured by conventional techniques to produce cells containing an antibody described herein or a fragment thereof. Thus, described herein are host cells containing a polynucleotide encoding containing an antibody described herein or fragments thereof, or a heavy or light chain thereof, or fragment thereof, or a single-chain antibody described herein, operably linked to a promoter for expression of such sequences in the host cell.

[0381] In certain embodiments, a host cell comprises a polynucleotide encoding the VH and VL of an antibody described herein. In another embodiment, a host cell comprises a vector comprising a polynucleotide encoding the VH and VL of an antibody described herein. In another embodiment, a host cell comprises a first polynucleotide encoding the VH of an antibody described herein, and a second polynucleotide encoding the VL of an antibody described herein. In anotherembodiment, a host cell comprises a first vector comprising a first polynucleotide encoding the VH of an antibody described herein, and a second vector comprising a second polynucleotide encoding the VL of an antibody described herein.

[0382] In specific embodiments, a heavy chain / heavy chain variable region expressed by a first cell is associated with a light chain / light chain variable region of a second cell to form an anti-LY6K (e.g., human LY6K) antibody described herein. In certain embodiments, described herein is a population of host cells comprising such first host cell and such second host cell.

[0383] In certain embodiments, described herein is a population of vectors comprising a first vector comprising a polynucleotide encoding a light chain / light chain variable region of an anti-LY6K (e.g., human LY6K) antibody described herein, and a second vector comprising a polynucleotide encoding a heavy chain / heavy chain variable region of an anti-LY6K (e.g, human LY6K) antibody described herein.

[0384] A variety of host-expression vector systems can be utilized to express antibody molecules described herein (see, e.g., U. S. Patent No. 5,807,715, which is herein incorporated by reference in its entirety). Such host-expression systems represent vehicles by which the coding sequences of interest can be produced and subsequently purified, but also represent cells which can, when transformed or transfected with the appropriate nucleotide coding sequences, express an antibody molecule described herein in situ. These include but are not limited to microorganisms such as bacteria e.g., E. coli and B. subtilis) transformed with, e.g., recombinant bacteriophage DNA, plasmid DNA or cosmid DNA expression vectors containing antibody coding sequences; yeast (e.g., Saccharomyces and Pichid) transformed with, e.g., recombinant yeast expression vectors containing antibody coding sequences; insect cell systems infected with, e.g., recombinant virus expression vectors (e.g., baculovirus) containing antibody coding sequences; plant cell systems (e.g., green algae such as Chlamydomonas reinhardtii) infected with, e.g., recombinant virus expression vectors (e.g., cauliflower mosaic virus, CaMV; tobacco mosaic virus, TMV) or transformed with, e.g., recombinant plasmid expression vectors (e.g., Ti plasmid) containing antibody coding sequences; or mammalian cell systems (e.g., COS (e.g., COS1 or COS), CHO, BHK, MDCK, HEK 293, NS0, PER. C6, VERO, CRL7O3O, HsS78Bst, HeLa, and NIH 3T3, HEK-293T, HepG2, SP210, Rl.1, B-W, L-M, BSC1, BSC40, YB / 20 and BMT10 cells) harboring, e.g., recombinant expression constructs containing promoters derived from the genome of mammalian cells (e.g., metallothionein promoter) or from mammalian viruses (e.g., the adenoviruslate promoter; the vaccinia virus 7.5K promoter). In a specific embodiment, cells for expressing antibodies described herein are Chinese hamster ovary (CHO) cells, for example CHO cells from the CHO GS System™ (Lonza). In certain embodiments, the heavy chain and / or light chain of an antibody produced by a CHO cell may have an N-terminal glutamine or glutamate residue replaced by pyroglutamate. In certain embodiments, cells for expressing antibodies described herein are human cells, e.g, human cell lines. In a specific embodiment, a mammalian expression vector is pOptiVEC™ or pcDNA3.3. In certain embodiments, bacterial cells such as Escherichia coli, or eukaryotic cells (e.g., mammalian cells), especially for the expression of whole recombinant antibody molecule, are used for the expression of a recombinant antibody molecule. For example, mammalian cells such as CHO cells, in conjunction with a vector such as the major intermediate early gene promoter element from human cytomegalovirus, are an effective expression system for antibodies (Foecking MK & Hofstetter H (1986) Gene 45: 101-5; and Cockett MI el al., (1990) Biotechnology 8(7): 662-7, each of which is herein incorporated by reference in its entirety). In certain embodiments, antibodies described herein are produced by CHO cells or NSO cells. In a specific embodiment, the expression of nucleotide sequences encoding antibodies described herein which specifically bind to LY6K (e.g., human LY6K) is regulated by a constitutive promoter, inducible promoter, or tissue specific promoter.

[0385] In bacterial systems, a number of expression vectors can be advantageously selected depending upon the use intended for the antibody molecule being expressed. For example, when a large quantity of such an antibody is to be produced, for the generation of pharmaceutical compositions of an antibody molecule, vectors which direct the expression of high levels of fusion protein products that are readily purified can be desirable. Such vectors include, but are not limited to, the E. coli expression vector pUR278 (Ruether U & Mueller-Hill B (1983) EMBO J 2: 1791-1794), in which the coding sequence can be ligated individually into the vector in frame with the lac Z coding region so that a fusion protein is produced; pIN vectors (Inouye S & Inouye M (1985) Nuc Acids Res 13: 3101-3109; Van Heeke G & Schuster SM (1989) J Biol Chem 24: 5503-5509); and the like, all of which are herein incorporated by reference in their entireties. For example, pGEX vectors can also be used to express foreign polypeptides as fusion proteins with glutathione 5-transferase (GST). In general, such fusion proteins are soluble and can easily be purified from lysed cells by adsorption and binding to matrix glutathione agarose beads followed by elution inthe presence of free glutathione. The pGEX vectors are designed to include thrombin or factor Xa protease cleavage sites so that the cloned target gene product can be released from the GST moiety.[003861 In an insect system, Autographa californica nuclear polyhedrosis virus (AcNPV), for example, can be used as a vector to express foreign genes. The virus grows in Spodoptera frugiperda cells. The coding sequence can be cloned individually into non-essential regions (for example the polyhedrin gene) of the virus and placed under control of an AcNPV promoter (for example the polyhedrin promoter).

[0387] In mammalian host cells, a number of viral-based expression systems can be utilized. In cases where an adenovirus is used as an expression vector, the coding sequence of interest can be ligated to an adenovirus transcription / translation control complex, e.g., the late promoter and tripartite leader sequence. This chimeric gene can then be inserted in the adenovirus genome by in vitro or in vivo recombination. Insertion in a non-essential region of the viral genome (e.g, region El or E3) will result in a recombinant virus that is viable and capable of expressing the molecule in infected hosts (see, e.g., Logan J & Shenk T (1984) PNAS 81(12): 3655-9, which is herein incorporated by reference in its entirety). Specific initiation signals can also be required for efficient translation of inserted coding sequences. These signals include the ATG initiation codon and adjacent sequences. Furthermore, the initiation codon must be in phase with the reading frame of the desired coding sequence to ensure translation of the entire insert. These exogenous translational control signals and initiation codons can be of a variety of origins, both natural and synthetic. The efficiency of expression can be enhanced by the inclusion of appropriate transcription enhancer elements, transcription terminators, etc. (see, e.g., Bitter G et al., (1987) Methods Enzymol. 153: 516-544, which is herein incorporated by reference in its entirety).

[0388] In addition, a host cell strain can be chosen which modulates the expression of the inserted sequences or modifies and processes the gene product in the specific fashion desired. Such modifications (e.g., glycosylation) and processing (e.g., cleavage) of protein products can be important for the function of the protein. Different host cells have characteristic and specific mechanisms for the post-translational processing and modification of proteins and gene products. Appropriate cell lines or host systems can be chosen to ensure the correct modification and processing of the foreign protein expressed. To this end, eukaryotic host cells which possess the cellular machinery for proper processing of the primary transcript, glycosylation, and phosphorylation of the gene product can be used. Such mammalian host cells include but are notlimited to CHO, VERO, BHK, Hela, MDCK, HEK 293, NTH 3T3, W138, BT483, Hs578T, HTB2, BT2O and T47D, NSO (a murine myeloma cell line that does not endogenously produce any immunoglobulin chains), CRL7O3O, COS (e.g., COS1 or COS), PER. C6, VERO, HsS78Bst, HEK-293T, HepG2, SP210, Rl.l, B-W, L-M, BSC1, BSC40, YB / 20, BMT10, and HsS78Bst cells. In certain embodiments, anti-LY6K (e.g., human LY6K) antibodies described herein are produced in mammalian cells, such as CHO cells.

[0389] In a specific embodiment, the antibodies described herein have reduced fucose content or no fucose content. Such antibodies can be produced using techniques known one skilled in the art. For example, the antibodies can be expressed in cells deficient or lacking the ability to fucosylate. In a specific example, cell lines with a knockout of both alleles of al, 6-fucosyltransferase can be used to produce antibodies with reduced fucose content. The Potelligent® system (Lonza) is an example of such a system that can be used to produce antibodies with reduced fucose content.

[0390] For long-term, high-yield production of recombinant proteins, stable expression cells can be generated. For example, cell lines which stably express an anti-LY6K (e.g., human LY6K) antibody described herein can be engineered. In specific embodiments, a cell described herein stably expresses a light chain / light chain variable region and a heavy chain / heavy chain variable region which associate to form an antigen-binding region or an antibody described herein.

[0391] In certain aspects, rather than using expression vectors which contain viral origins of replication, host cells can be transformed with DNA controlled by appropriate expression control elements (e.g., promoter, enhancer, sequences, transcription terminators, polyadenylation sites, etc.), and a selectable marker. Following the introduction of the foreign DNA / polynucleotide, engineered cells can be allowed to grow for 1-2 days in an enriched media, and then are switched to a selective media. The selectable marker in the recombinant plasmid confers resistance to the selection and allows cells to stably integrate the plasmid into their chromosomes and grow to form foci which in turn can be cloned and expanded into cell lines. This method can advantageously be used to engineer cell lines which express an anti-LY6K (e.g., human LY6K) described herein or a fragment thereof. Such engineered cell lines can be particularly useful in screening and evaluation of compositions that interact directly or indirectly with the antibody molecule.

[0392] A number of selection systems can be used, including but not limited to the herpes simplex virus thymidine kinase (Wigler Metal., (1977) Cell 11(1): 223-32), hypoxanthineguaninephosphoribosyltransferase (SzybalskaEH & Szybalski W (1962) PNAS 48(12): 2026-2034), and adenine phosphoribosyltransferase (Lowy I et al., (1980) Cell 22(3): 817-23) genes in tk-, hgprt-or aprt-cells, respectively, all of which are herein incorporated by reference in their entireties. Also, antimetabolite resistance can be used as the basis of selection for the following genes: dhfr, which confers resistance to methotrexate (Wigler M et al., (1980) PNAS 77(6): 3567-70; O’Hare K et al., (1981) PNAS 78: 1527-31); gpt, which confers resistance to mycophenolic acid (Mulligan RC & Berg P (1981) PNAS 78(4): 2072-6); neo, which confers resistance to the aminoglycoside G-418 (Wu GY & Wu CH (1991) Biotherapy 3: 87-95; Tolstoshev P (1993) Ann Rev Pharmacol Toxicol 32: 573-596; Mulligan RC (1993) Science 260: 926-932; and Morgan RA & Anderson WF (1993) Ann Rev Biochem 62: 191-217; Nabel GJ & Feigner PL (1993) Trends Biotechnol 11(5): 211-5); and hygro, which confers resistance to hygromycin (Santerre RF etal., (1984) Gene 30(1-3): 147-56), all of which are herein incorporated by reference in their entireties. Methods commonly known in the art of recombinant DNA technology can be routinely applied to select the desired recombinant clone and such methods are described, for example, in Ausubel FM et al., (eds.), Current Protocols in Molecular Biology, John Wiley & Sons, NY (1993); Kriegler M, Gene Transfer and Expression, A Laboratory Manual, Stockton Press, NY (1990); and in Chapters 12 and 13, Dracopoli NC et al, (eds.), Current Protocols in Human Genetics, John Wiley & Sons, NY (1994); Colbere-Garapin F et al., (1981) J Mol Biol 150: 1-14, all of which are herein incorporated by reference in their entireties.

[0393] The expression levels of an antibody molecule can be increased by vector amplification (for a review, see Bebbington CR & Hentschel CCG, The use of vectors based on gene amplification for the expression of cloned genes in mammalian cells in DNA cloning, Vol. 3 (Academic Press, New York, 1987), which is herein incorporated by reference in its entirety). When a marker in the vector system is amplifiable, an increase in the level of inhibitor present in culture of the host cell will increase the number of copies of the marker gene. Since the amplified region is associated with the gene of interest, production of the protein will also increase (Crouse GF etal., (1983) Mol Cell Biol 3: 257-66, which is herein incorporated by reference in its entirety).

[0394] The host cell can be co-transfected with two or more expression vectors described herein, the first vector encoding a heavy chain derived polypeptide and the second vector encoding a light chain derived polypeptide. The two vectors can contain identical selectable markers which enable equal expression of heavy and light chain polypeptides. The host cells can be co-transfectedwith different amounts of the two or more expression vectors. For example, host cells can be transfected with any one of the following ratios of a first expression vector and a second expression vector: about 1:1, 1:2, 1:3, 1:4, 1:5, 1:6, 1:7, 1:8, 1:9, 1:10, 1:12, 1:15, 1:20, 1:25, 1:30, 1:35, 1:40, 1:45, or 1:50.

[0395] Alternatively, a single vector can be used which encodes, and is capable of expressing, both heavy and light chain polypeptides. In such situations, the light chain should be placed before the heavy chain to avoid an excess of toxic free heavy chain (Proudfoot NJ (1986) Nature 322: 562-565; and Kohler G (1980) PNAS 77: 2197-2199, each of which is herein incorporated by reference in its entirety). The coding sequences for the heavy and light chains can comprise cDNA or genomic DNA. The expression vector can be monocistronic or multicistronic. A multi ci stronic nucleic acid construct can encode 2, 3, 4, 5, 6, 7, 8, 9, 10, or more genes / nucleotide sequences, or in the range of 2-5, 5-10, or 10-20 genes / nucleotide sequences. For example, a bicistronic nucleic acid construct can comprise, in the following order, a promoter, a first gene e.g., heavy chain of an antibody described herein), and a second gene and (e.g., light chain of an antibody described herein). In such an expression vector, the transcription of both genes can be driven by the promoter, whereas the translation of the mRNA from the first gene can be by a cap-dependent scanning mechanism and the translation of the mRNA from the second gene can be by a cap-independent mechanism, e.g., by an IRES.

[0396] Once an antibody molecule described herein has been produced by recombinant expression, it can be purified by any method known in the art for purification of an immunoglobulin molecule, for example, by chromatography (e.g., ion exchange, affinity, particularly by affinity for the specific antigen after Protein A, and sizing column chromatography), centrifugation, differential solubility, or by any other standard technique for the purification of proteins. Further, the antibodies described herein can be fused to heterologous polypeptide sequences described herein or otherwise known in the art to facilitate purification.

[0397] In specific embodiments, an antibody described herein is isolated or purified. In certain embodiments, an isolated antibody is one that is substantially free of other antibodies with different antigenic specificities than the isolated antibody. For example, in certain embodiments, a preparation of an antibody described herein is substantially free of cellular material and / or chemical precursors. The language “substantially free of cellular material” includes preparations of an antibody in which the antibody is separated from cellular components of the cells from whichit is isolated or recombinantly produced. Thus, an antibody that is substantially free of cellular material includes preparations of antibody having less than about 30%, 20%, 10%, 5%, 2%, 1%, 0.5%, or 0.1% (by dry weight) of heterologous protein (also referred to herein as a “contaminating protein”) and / or variants of an antibody, for example, different post-translational modified forms of an antibody or other different versions of an antibody (e.., antibody fragments). When the antibody is recombinantly produced, it is also generally substantially free of culture medium, i...

Claims

WHAT IS CLAIMED:

1. An antibody-drug conjugate (ADC) according to general structure (1):AB-[(L6)b-{Z-L-D}x]y(1)wherein:AB is an antibody that specifically binds human LY6K;L is a linker that links Z to D;Z is a connecting group;L6is -GlcNAc(Fuc)w-(G)j-S-(L7)w’-, wherein G is a monosaccharide, j is an integer in the range of 0 - 10, S is a sugar or a sugar derivative, GlcNAc is N-acetylglucosamine and Fuc is fucose, w is 0 or 1, w’ is 0, 1 or 2 and L7is -N(H)C(O)CH2-, -N(H)C(O)CF2- or -CH2-;D is selected from the group consisting of taxanes, anthracyclines, camptothecins, epothilones, mytomycins, combretastatins, vinca alkaloids, tubulysins, enediynes, amanitins, duocarmycins, maytansinoids, amatoxins, dolastatins, auristatins, eribulins, BCL-XL inhibitors, hemiasterlins, KSP inhibitors, radioisotopes, TLR agonists, indolinobenzodiazepine dimers or pyrrolobenzodiazepine dimers (PBDs), and analogues or prodrugs thereof;b is 0 or 1;x is 1 or 2; andy is 1, 2, 3 or 4.

2. The ADC of claim 1, wherein linker L has the structure-(L1)n-(L2)0-(L3)p-(L4)q-wherein:L1, L2, L3and L4are each individually linkers that together link Z to D;n, o, p and q are each individually 0 or 1, provided that n + o + p + q= l, 2, 3 or 4, optionally wherein n = o = p = 1;and wherein:(a) linker L1is represented by:-(W)k-(A)d-(B)e-(A)f-(B)g-(C(O))g-, or-(W)k-(A)d-(B)e-(A)f-(C(O))g-N*[-(A)d’-(B)e’-(A)f’-(C(O))g— ]2,wherein:d and d’ are individually 0 or 1;e and e’ are individually an integer in the range 1 - 10;f and f are individually 0, or 1;g and g’ are individually an integer in the range 0 - 10;k = 0 or 1 with the proviso that if k = 1 then d = 0;A is a sulfamide group according to structure (23)wherein a = 0 or 1, and R13is selected from the group consisting of hydrogen, Ci - C24 alkyl groups, C3 - C24 cycloalkyl groups, C2 - C24 (hetero)aryl groups, C3 - C24 alkyl(hetero)aryl groups and C3 - C24 (hetero)arylalkyl groups, the Ci - C24 alkyl groups, C3 - C24 cycloalkyl groups, C2 - C24 (hetero)aryl groups, C3 - C24 alkyl(hetero)aryl groups and C3 - C24 (hetero)aryl alkyl groups optionally substituted and optionally interrupted by one or more heteroatoms selected from O, S and NR14wherein R14is independently selected from the group consisting of hydrogen and Ci - C4 alkyl groups, or R13is D connected to N via a spacer moiety, optionally wherein the spacer moiety is -(B)g-(C(O))g-(L2)o-(L3)p-(L4)q-;W is -OC(O)-, -C(O)O- -C(O)NH- -NHC(O)-, -OC(O)NH- -NHC(O)O- -C(O)(CH2)mC(O)-, -C(O)(CH2)niC(O)NH- or -(4-Ph)CH2NHC(O)(CH2)mC(O)NH- wherein m is an integer in the range 0 - 10;B is a -CH2-CH2-O- or a -O-CH2-CH2- moiety, or (B)eis a -(CH2-CH2-O)ei-CH2- CH2- moiety, wherein el is an integer in the range 1 - 10;N* is a branching nitrogen atom, to which two instances of-(A)d-(B)e-(A)f-(C(O))g’- are connected and both (C(O))g’ moieties are connected to -(L2)o-(L3)p-(L4)q-D, wherein L2, L3, L4, o, p, q and D are each selected individually;and / or(b) linker L2is a peptide spacer, optionally wherein linker L2comprises 1 - 5 amino acids, wherein linker L2is a dipeptide, tripeptide or tetrapeptide spacer, or wherein linker L2is represented by general structure (L3):wherein R17= CH3or CH2CH2CH2NHC(O)NH2;and / or(c) linker L3is a self-immolative spacer, optionally wherein linker L3is a paraaminobenzyloxycarbonyl (PABC) derivative according to structure (L4).wherein R21is H, R26or C(O) R26, wherein R26is Ci - C24 (hetero)alkyl groups, C3 - C10 (hetero)cycloalkyl groups, C2 - C10 (hetero)aryl groups, C3 - C10 alkyl(hetero)aryl groups and C3 - C10 (hetero)arylalkyl groups, which are optionally substituted and optionally interrupted by one or more heteroatoms selected from O, S and NR28, wherein R28is independently selected from the group consisting of hydrogen and Ci - C4 alkyl groups,optionally wherein R21is H or C(O)R26, wherein R26= 4-methyl-piperazine or morpholine;and / or(d) linker L4is an aminoalkanoic acid spacer according to the structure - NR22-(Cz-alkylene)- C(O)-, wherein x is an integer in the range 1 -20 and R22is H or Ci - C4 alkyl; or linker L4is a an ethyleneglycol spacer according to the structure -NR22-(CH2-CH2-O)e6- (CH2)e7-C(O)-, wherein e6 is an integer in the range 1 - 10, e7 is an integer in the range 1 - 3 and R22is H or Ci - C4 alkyl; orlinker L4is a diamine spacer according to the structure -NR22-(Cz-alkylene)-NR22- (C(O))h-, wherein h is 0 or 1, x is an integer in the range 1 - 10 and R22is H or Ci C4 alkyl.

3. The ADC of claim 1 or 2, wherein b = 1 and conjugation is via the glycan of the antibody, optionally wherein j = 0.

4. The ADC of any one of the preceding claims, wherein L-D has a structure selected from the group consisting of (X) - (XIII):wherein the wavy line indicates the connection to Z, and L2, L4, o and q are as defined in claim 2.

5. The ADC of claim 4, wherein L-D has a structure selected from the group consisting of (Xa), (Xlb), (Xllg), (XHIe) and (Xllh):wherein the wavy line indicates the connection to Z, and R22is as defined in claim 38.

6. The ADC of any one of the preceding claims, wherein Z comprises a cyclooctenyl groupwherein:the bond labelled with * is connected to S and the wavy bond labelled with ** is connected to L;each R15is independently selected from the group consisting of hydrogen, halogen, - OR16, -NO2, -CN, -S(O)2R16, -S(O)3(-), CI - C24 alkyl groups, C5 - C24 (hetero)aryl groups, C7 - C24 alkyl(hetero)aryl groups and C7 - C24 (hetero)aryl alkyl groups and wherein the alkyl groups, (hetero)aryl groups, alkyl(hetero)aryl groups and (hetero)arylalkyl groups are optionally substituted, wherein two substituents R15may be linked together to form an optionally substituted annulated cycloalkyl or an optionally substituted annulated (hetero)arene substituent, and wherein R16is independently selected from the group consisting of hydrogen,halogen, Ci - C24 alkyl groups, Ce - C24 (hetero)aryl groups, C7 - C24 alkyl(hetero)aryl groups and C7 - C24 (hetero)aryl alkyl groups;each R18is independently selected from the group consisting of hydrogen, halogen, Ci - C24 alkyl groups, Ce - C24 (hetero)aryl groups, C7 - C24 alkyl(hetero)aryl groups and C7 - C24 (hetero)arylalkyl groups;R19is selected from the group consisting of hydrogen, halogen, Ci - C24 alkyl groups, Ce - C24 (hetero)aryl groups, C7 - C24 alkyl(hetero)aryl groups and C7 - C24 (hetero)arylalkyl groups, the alkyl groups optionally being interrupted by one of more hetero-atoms selected from the group consisting of O, N and S, wherein the alkyl groups, (hetero)aryl groups, alkyl(hetero)aryl groups and (hetero)arylalkyl groups are independently optionally substituted, or R19is a second occurrence of Z (or Q) or D connected via a spacer moiety; and1 is an integer in the range 0 to 10.

7. An antibody-drug conjugate (ADC) according to general structure (1):AB-[(L6)b-{Z-L-D}x]y(1)wherein the ADC is obtained by a process comprising:(i) contacting an antibody that specifically binds human LY6K comprising y core N-acetylglucosamine (GlcNAc) moi eties, wherein y = 1, 2, 3 or 4, with a compound of the formula S(F)X-P in the presence of a catalyst, wherein S(F)Xis a sugar derivative comprising x reactive groups F capable of reacting with a reactive group Q, x is 1 or 2 and P is a nucleoside mono- or diphosphate, and wherein the catalyst is capable of transferring the S(F)Xmoiety to the core-GlcNAc moiety, to obtain a modified antibody according to Formula (26):AB-[GlcNAc(Fuc)w-S {F }x]y(26)whereinAB is the antibody that specifically binds human LY6K;Fuc is fucose; andw is 0 or 1; and(ii) reacting the modified antibody with a compound according to structure (2):Q-L-D(2)wherein:Q is a reactive moiety;is a linker that links Z to D; andD is selected from the group consisting of taxanes, anthracyclines, camptothecins, epothilones, mytomycins, combretastatins, vinca alkaloids, tubulysins, enediynes, amanitins, duocarmycins, maytansinoids, amatoxins, dolastatins, auristatins, eribulins, BCL-XL inhibitors, hemiasterlins, KSP inhibitors, radioisotopes, TLR agonists, indolinobenzodiazepine dimers or pyrrolobenzodiazepine dimers (PBDs), and analogues or prodrugs thereof;to obtain the ADC.

8. The ADC of any one of the preceding claims, wherein the antibody that specifically binds human LY6K comprises: a VH comprising the CDRH1, CDRH2, and CDRH3 amino acid sequences of the VH amino acid sequence set forth in any one of SEQ ID NOs: 1-34; and a VL comprising the CDRL1, CDRL2, and CDRL3 amino acid sequences of the VL amino acid sequence set forth in any one of SEQ ID NOs: 35-68.

9. The ADC of claim 8, wherein the VH amino acid sequence and VL amino acid sequence are as set forth in: SEQ ID NOs: 1 and 35, 2 and 36, 3 and 37, 4 and 38, 5 and 39, 6 and 40, 7 and 41, 8 and 42, 9 and 43, 10 and 44, 11 and 45, 12 and 46, 13 and 47, 14 and 48, 15 and 49, 16 and 50, 17 and 51, 18 and 52, 19 and 53, 20 and 54, 21 and 55, 22 and 56, 23 and 57, 24 and 58, 25 and 59, 26 and 60, 27 and 61, 28 and 62, 29 and 63, 30 and 64, 31 and 65, 32 and 66, 33 and 67, or 34 and 68, respectively.

10. The ADC of claim 8 or 9, wherein the antibody comprises the CDRH1, CDRH2, and CDRH3 amino acid sequences set forth in: SEQ ID NOs: 69, 90, and 124; 70, 91, and 125; 71, 92, and 126; 72, 93, and 127; 69, 94, and 124; 73, 95, and 128; 74, 96, and 129; 75, 97, and 130; 74, 98, and 131; 76, 99, and 132; 77, 100, and 133; 78, 101, and 134; 79, 102, and 135; 74, 103, and 136; 80, 104, and 137; 74, 105, and 138; 74, 106, and 139; 81, 107, and 140; 81, 108, and 141; 82, 109, and 142; 83, 110, and 143; 84, 99, and 144; 85, 111, and 145; 81, 112, and 146; 86, 113, and147; 87, 114, and 148; 74, 115, and 149; 74, 116, and 150; 74, 117, and 151; 88, 118, and 152; 89, 119, and 153; 81, 120, and 154; 74, 121, and 155; 85, 122, and 156; or 86, 123, and 157, respectively.

11. The ADC of any one of claims 8-10, wherein the antibody comprises the CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in: SEQ ID NO: 158, KIS, and SEQ ID NO: 198; SEQ ID NO: 159, WAS, and SEQ ID NO: 199; SEQ ID NO: 160, STT, and SEQ ID NO: 200; SEQ ID NO: 161, WAS, and SEQ ID NO: 201; SEQ ID NO: 162, AAS, and SEQ ID NO: 202; SEQ ID NOs: 163, 185, and 203; SEQ ID NOs: 164, 185, and 204; SEQ ID NOs: 165, 185, and 205; SEQ ID NOs: 166, 186, and 206; SEQ ID NOs: 167, 187, and 207; SEQ ID NOs: 168, 188, and 208; SEQ ID NOs: 169, 186, and 209; SEQ ID NOs: 170, 185, and 198; SEQ ID NOs: 169, 189, and 210; SEQ ID NOs: 170, 185, and 211; SEQ ID NOs: 171, 187, and 212; SEQ ID NOs: 172, 187, and 213; SEQ ID NOs: 173, 190, and 214; SEQ ID NOs: 174, 191, and 215; SEQ ID NOs: 175, 186, and 216; SEQ ID NOs: 176, 186, and 217; SEQ ID NOs: 170, 192, and 198; SEQ ID NOs: 171, 193, and 218; SEQ ID NOs: 177, 187, and 219; SEQ ID NOs: 178, 189, and 220; SEQ ID NOs: 179, 187, and 221; SEQ ID NOs: 165, 185, and 222; SEQ ID NOs: 180, 187, and 223; SEQ ID NOs: 181, 194, and 224; SEQ ID NOs: 182, 195, and 199; SEQ ID NOs: 172, 187, and 225; SEQ ID NOs: 183, 196, and 226; or SEQ ID NOs: 184, 197, and 227, respectively.

12. The ADC of any one of claims 8-11, wherein the antibody comprises the CDRH1, CDRH2, CDRH3, CDRL1, CDRL2, and CDRL3 amino acid sequences set forth in: SEQ ID NOs: 69, 90, 124, 158, KIS, and SEQ ID NO: 198; SEQ ID NOs: 70, 91, 125, 159, WAS, and SEQ ID NO: 199; SEQ ID NOs: 71, 92, 126, 160, STT, and SEQ ID NO: 200; SEQ ID NOs: 72, 93, 127, 161, WAS, and SEQ ID NO: 201; SEQ ID NOs: 69, 94, 124, 158, KIS, and SEQ ID NO: 198; SEQ ID NOs: 73, 95, 128, 162, AAS, and SEQ ID NO: 202; SEQ ID NOs: 74, 96, 129, 163, 185, and 203; SEQ ID NOs: 75, 97, 130, 164, 185, and 204; SEQ ID NOs: 74, 98, 131, 165, 185, and 205; SEQ ID NOs: 76, 99, 132, 166, 186, and 206; SEQ ID NOs: 77, 100, 133, 167, 187, and 207; SEQ ID NOs: 78, 101, 134, 168, 188, and 208; SEQ ID NOs: 79, 102, 135, 169, 186, and 209; SEQ ID NOs: 74, 103, 136, 170, 185, and 198; SEQ ID NOs: 80, 104, 137, 169, 189, and 210; SEQ ID NOs: 74, 105, 138, 170, 185, and 211; SEQ ID NOs: 74, 106, 139, 171, 187, and 212; SEQ ID NOs: 81, 107, 140, 172, 187, and 213; SEQ ID NOs: 81, 108, 141, 173, 190, and 214; SEQ ID NOs: 82,109, 142, 174, 191, and 215; SEQ IDNOs: 83, 110, 143, 175, 186, and 216; SEQ IDNOs: 84, 99, 144, 176, 186, and 217; SEQ ID NOs: 85, 111, 145, 170, 192, and 198; SEQ ID NOs: 81, 112, 146, 171, 193, and 218; SEQ ID NOs: 86, 113, 147, 177, 187, and 219; SEQ ID NOs: 87, 114, 148, 178, 189, and 220; SEQ ID NOs: 74, 115, 149, 179, 187, and 221; SEQ ID NOs: 74, 116, 150, 165, 185, and 222; SEQ ID NOs: 74, 117, 151, 180, 187, and 223; SEQ ID NOs: 88, 118, 152, 181, 194, and 224; SEQ ID NOs: 89, 119, 153, 182, 195, and 199; SEQ ID NOs: 81, 120, 154, 172, 187, and 225; SEQ ID NOs: 74, 121, 155, 165, 185, and 222; SEQ ID NOs: 85, 122, 156, 183, 196, and 226; or SEQ ID NOs: 86, 123, 157, 184, 197, and 227, respectively.

13. The ADC of any one of claims 8-12, wherein the antibody comprises the VH amino acid sequence set forth in any one of SEQ ID NOs: 1-34.

14. The ADC of any one of claims 8-13, wherein the antibody comprises a heavy chain constant region, optionally selected from the group consisting of human IgGi, IgG2, IgGs, IgG4, IgAi, and IgA215. The ADC of any one of claims 8-14, wherein the antibody comprises a heavy chain constant region that is a variant of a wild-type heavy chain constant region, wherein the variant heavy chain constant region binds to an FcyR with lower affinity than the wild-type heavy chain constant region binds to the FcyR.

16. The ADC of any one of claims 8-15, wherein the antibody comprises a heavy chain constant region comprising the amino acid sequence of SEQ ID NO: 228 or 229.

17. The ADC of any one of claims 8-16, wherein the antibody comprises the VL amino acid sequence set forth in any one of SEQ ID NOs: 35-68.

18. The ADC of any one of claims 8-17, wherein the antibody comprises a light chain constant region comprising the amino acid sequence of SEQ ID NO: 230 or 231.

19. The ADC of any one of claims 8-18, wherein the VH and VL comprise the amino acid sequences set forth in: SEQ ID NOs: 1 and 35, 2 and 36, 3 and 37, 4 and 38, 5 and 39, 6 and 40, 7 and 41, 8 and 42, 9 and 43, 10 and 44, 11 and 45, 12 and 46, 13 and 47, 14 and 48, 15 and 49, 16 and 50, 17 and 51, 18 and 52, 19 and 53, 20 and 54, 21 and 55, 22 and 56, 23 and 57, 24 and 58, 25 and 59, 26 and 60, 27 and 61, 28 and 62, 29 and 63, 30 and 64, 31 and 65, 32 and 66, 33 and 67, or 34 and 68, respectively, respectively.

20. The ADC of any one of claims 8-19, wherein the antibody specifically binds to an amino acid sequence selected from the group consisting of SEQ ID NOs: 233-238.

21. The ADC of any one of claims 8-19, wherein the antibody specifically binds to the amino acid sequence of SEQ ID NOs: 233 and 239.

22. The ADC of any one of claims 8-19, wherein the antibody specifically binds to the amino acid sequence of SEQ ID NOs: 234 and 240.

23. The ADC of any one of claims 8-19, wherein the antibody specifically binds to the amino acid sequence of SEQ ID NOs: 237 and 247.

24. The ADC of any one of the preceding claims, wherein D is selected from the group consisting of anthracyclines, camptothecins, maytansinoids, enediynes, auristatins and pyrrolobenzodiazepine dimers.

25. The ADC of any one of claims 1-24, wherein D is an auristatin selected from the group consisting of MMAD, MMAE, MMAF and PF -06380101.

26. The ADC of claim 25, wherein D is MMAE or PF-06380101.

27. The ADC of claim 25 or 26, wherein D is MMAE.

28. The ADC of any one of claims 1 -24, wherein D is a camptothecin selected from the group consisting of topotecan, silatecan, cositecan, exatecan, exatecan-S, DXd, SN-38, lurtotecan, gimatecan, belotecan, rubitecan, AMDCPT, and G-AMDCPT.

29. The ADC of claim 28, wherein D is exatecan or DXd.

30. The ADC of claim 28 or 29, wherein D is exatecan.

31. The ADC of any one of claims 1-24, wherein D is an enediyne selected from the group consisting of calicheamicins, esperamicins, shishijimicins, and namenamicins.

32. The ADC of claim 31, wherein D is calicheamicin.

33. A composition comprising the ADC of any one of the preceding claims and a pharmaceutically acceptable carrier or excipient.

34. A method of treating cancer in a subject, the method comprising administering to the subject an effective amount of: the ADC of any one of claims 1-32 or the composition of claim 33.

35. The method of claim 34, further comprising administering an additional therapeutic agent to the subject.

36. The method of claim 35, wherein the additional therapeutic agent is a chemotherapeutic, a radiotherapeutic, or a checkpoint targeting agent.

37. The method of claim 36, wherein the checkpoint targeting agent is selected from the group consisting of an antagonist anti-PD-1 antibody, an antagonist anti-PD-Ll antibody, an antagonist anti-PD-L2 antibody, an antagonist anti-CTLA-4 antibody, an antagonist anti-TIM-3 antibody, an antagonist anti-LAG-3 antibody, an antagonist anti-VISTA antibody, an antagonist anti-TIGIT antibody, an antagonist anti-CD96 antibody, an antagonist anti-CEACAMl antibody, an agonist anti-CD137 antibody, an agonist anti-GITR antibody, and an agonist anti-OX40 antibody.

38. Use of the ADC of any one of claims 1-32 or the composition of claim 33 for the manufacture of a medicament for treating cancer.

39. The ADC of any one of claims 1-32 or the composition of claim 33 for use in medicine.

40. The ADC of any one of claims 1-32 or the composition of claim 33 for use in treating cancer in a subject.

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